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Central cholinergic system and endocannabinoid, anandamide exhibits anti-compulsive-like behavior in mice. However, the role of the central cholinergic system in the anandamide-induced anti-compulsive-like behavior is still unexplored. Therefore, the present study assessed the role of central cholinergic transmission in the anandamide-induced anti-compulsive activity using a marble-burying behavior (MBB) model in mice. The modulation in the anandamide-induced effect on MBB was evaluated using mice with altered central cholinergic transmission achieved by pretreatment (i.c.v.) with various cholinergic agents like acetylcholine (ACh), acetylcholinesterase inhibitor (AChEI), neostigmine, nicotine, mAChR antagonist, atropine, and nAChR antagonist, mecamylamine. The influence of anandamide treatment on the brain AChE activity was also evaluated. The results revealed that i.c.v. injection of anandamide (10, 20 µg/mouse, i.c.v.) dose-dependently reduced MBB in mice. Moreover, anandamide in all the tested doses inhibited the brain AChE activity indicating the role of an enhanced central cholinergic transmission in its anti-compulsive-like effect . Furthermore, the anti-compulsive-like effect of anandamide (20 µg/mouse, i.c.v.) was found to be enhanced in mice centrally pre-treated with, ACh (0.1 µg/mouse, i.c.v.) or AChEI, neostigmine (0.3 µg/mouse, i.c.v.). In addition, the anandamide-induced anti-compulsive-like effect was significantly increased in mice pre-treated with a low dose of nicotine (0.1 µg/mouse, i.c.v.) while, it was attenuated by the higher dose of nicotine (2 µg/mouse, i.c.v.). On the other hand, the anandamide (20 µg/mouse, i.c.v.) induced anti-compulsive-like effect was found to be diminished in mice pre-treated with mAChR antagonist, atropine (0.1, 0.5 µg/mouse, i.c.v.) and pre-injection of nAChR antagonist, mecamylamine (0.1, 0.5 µg/mouse, i.c.v.) potentiated the anandamide induced anti-compulsive-like response in mice. Thus, the present investigation delineates the modulatory role of an enhanced central cholinergic transmission in the anandamide-induced anti-compulsive-like behavior in mice by inhibition of brain AChE or via muscarinic and nicotinic receptors mediated mechanism., Competing Interests: Declaration of Competing Interest I the undersigned on the behalf of all authors certifies that the manuscripts entitled “Central cholinergic transmission modulates endocannabinoid-induced marble-burying behavior in mice” submitted to the journal behavioural brain research declares no conflict of interest., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Cholinergic dysfunction in the hippocampus causes memory impairment, and degradation of the forebrain cholinergic system has been implicated in several neurological disorders. One such disorder, Alzheimer's Disease (AD) is associated with the abnormal expression of various proteins including matrix metalloproteinase-9 (MMP-9), an enzyme known to regulate hippocampus-dependent memory. Memory involves several stages including acquisition, consolidation, and retrieval, but the neurobiological correlates of retrieval have been studied much less than other stages of memory. We sought to investigate the potential relationship between cholinergic signaling and hippocampal MMP-9 expression and the involvement of each in spatial memory retrieval. We trained rats in the water maze until the task was well-learned, then, seven days later, we allowed some to retrieve the memory after an intracerebroventricular injection of scopolamine or vehicle. Western blot analysis of hippocampal tissue shows elevated levels of a truncated form of MMP-9 associated with spatial memory retrieval. Additionally, our results indicate that centrally administered scopolamine both impairs spatial memory retrieval and prevents retrieval-induced elevations in MMP-9. These findings provide evidence for a potential link between cholinergic dysregulation and abnormal MMP-9 levels seen in the brains of AD patients. An important, yet unresolved question is whether MMP-9 serves to support memory retrieval itself or if it is involved in maintaining the ongoing stability of a retrieved memory., Competing Interests: Conflict of Interest Statement The authors have no conflicts of interest to declare., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Evidence has demonstrated the hippocampal cholinergic system and the mammalian target of rapamycin (mTOR) participation during the memory formation of aversive events. This study assessed the role of these systems in the hippocampus for the extinction memory process by submitting male Wistar rats to fear-motivated step-down inhibitory avoidance (IA). The post-extinction session administration of the nicotinic and muscarinic cholinergic receptor antagonists, mecamylamine and scopolamine, respectively, both at doses of 2 µg/µl/side, and rapamycin, an mTOR inhibitor (0.02 µg/µl/side), into the CA1 region of the dorsal hippocampus, impaired the IA extinction memory. Furthermore, the nicotinic and muscarinic cholinergic receptor agonists, nicotine and muscarine, respectively, had a dose-dependent effect on the IA extinction memory when administered intra-CA1, immediately after the extinction session. Nicotine (0.6 µg/µl/side) and muscarine (0.02 µg/µl/side), respectively, had no effect, while the higher doses (6 and 2 µg/µl/side, respectively) impaired the IA extinction memory. Interestingly, the co-administration of muscarine at the lower dose blocked the impairment that was induced by rapamycin. This effect was not observed when nicotine at the lower dose was co-administered. These results have demonstrated the participation of the cholinergic receptors and mTOR in the hippocampus for IA extinction, and that the cholinergic agonists had a dose-dependent effect on the IA extinction memory. This study provides insights related to the behavioural aspects and the neurobiological properties underlying the early stage of fear-motivated IA extinction memory consolidation and suggests that there is hippocampal muscarinic receptor participation independent of mTOR in this memory process., Competing Interests: Conflict of Interest The authors declare no conflicts of interest., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Cognitive flexibility facilitates adaptions to a changing environment in humans and animals and can be assessed with the attentional set shifting task (ASST). In various learning paradigms for laboratory rodents, the partial NMDA receptor agonist D-cycloserine has been found to have pro-cognitive effects. However, D-cycloserine has not yet been investigated for its effects on cognitive flexibility. The aim of the present study was to determine whether D-cycloserine is able to improve cognitive flexibility measured by the ASST in rats. Rats were first pre-treated with the muscarinic antagonist scopolamine (0.5 mg/kg) before the D-cycloserine administrations (20 mg/kg) to induce deficits in ASST performance. Our findings showed impaired ASST performance after scopolamine administration with significant effects on reversal phases and extra-dimensional shift. D-cycloserine treatment selectively improved the performance in the extra-dimensional shift and the last reversal phase, where scopolamine effects were most pronounced. These findings suggest that D-cycloserine can rescue deficits in cognitive flexibility., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Muscarinic acetylcholine receptors (mAChRs) play an important role in many brain functions. Our previous study revealed that the injection of mAChRs antagonist scopolamine into the intermediate medial mesopallium (IMM) region, which is critical for filial imprinting, impairs memory formation. In avian brains, four mAChR subtypes have been identified (M 2 , M 3 , M 4 and M 5 ). M 3 and M 5 receptors increase the excitability of neurons, whereas M 2 and M 4 receptors reduce the excitability. Because the scopolamine blocks all subtypes, the previous study did not identify which subtype contributes to the memory formation. By injecting several types of mAChR antagonists into the IMM, in this study we determined which mAChR subtype plays a critical role in imprinting. First, the effects of antagonists on the excitatory receptor subtypes M 3 and M 5 were examined. Injection of the M 3 antagonist (DAU5884) at 20 mM or the M 5 antagonist (ML381) at 2 mM impaired imprinting. Considering the pKi value of DAU5884, the impairment seems to be caused by DAU5884 binding to M 3 and/or M 4 receptors. Second, the effect of antagonists on the inhibitory receptor subtype M 2 was examined. The results showed that the M 2 antagonist (AQ-RA741) impaired imprinting at a concentration of 20 mM. Considering the pKi value of AQ-RA741, the impairment seems to be caused by AQ-RA741 binding to M 2 and/or M 4. The findings of this study suggests that the excitatory receptor subtypes M 3 and M 5 and the inhibitory receptor subtype M 2 and/or M 4 cooperate to achieve the appropriate balance of acetylcholine signaling to execute imprinting., (Copyright © 2022 Elsevier B.V. All rights reserved.)

The striatal beat frequency model assumes that striatal medium spiny neurons encode duration via synaptic plasticity. Muscarinic 1 (M1) cholinergic receptors as well as dopamine and glutamate receptors are important for neural plasticity in the dorsal striatum. Therefore, we investigated the effect of inhibiting these receptors on the formation of duration memory. After sufficient training in a peak interval (PI)-20-s procedure, rats were administered a single or mixed infusion of a selective antagonist for the dopamine D1 receptor (SCH23390, 0.5 µg per side), N-methyl-D-aspartic acid (NMDA)-type glutamate receptor (D-AP5, 3 µg), or M1 receptor (pirenzepine, 10 µg) bilaterally in the dorsal striatum, immediately before initiating a PI-40 s session (shift session). The next day, the rats were tested for new duration memory (40 s) in a session in which no lever presses were reinforced (test session). In the shift session, the performance was comparable irrespective of the drug injected. However, in the test session, the mean peak time (an index of duration memory) of the M1 + NMDA co-blockade group, but not of the D1 + NMDA co-blockade group, was lower than that of the control group (Experiments 1 and 2). In Experiment 3, the effect of the co-blockade of M1 and NMDA receptors was replicated. Moreover, sole blockade of M1 receptors induced the same effect as M1 and NMDA blockade. These results suggest that in the dorsal striatum, the M1 receptor, but not the D1 or NMDA receptors, is involved in the consolidation of duration memory., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Laboratory rats repeatedly exposed to an open field (OF) apparatus display increasingly high levels of grooming -especially that characterized by long and complex sequences- which has been taken as an additional index of novelty habituation. We hypothesized that disrupting such a learning process by administering an amnesic drug as the antimuscarinic scopolamine (SCP) could delay the appearance of more complex grooming subtypes. Thus, rats were pretreated either with SCP (15 mg/kg or 30 mg/kg) or vehicle (VEH) upon four one-day apart OF (OF1-4). On a fifth assessment, all rats received VEH to analyze the likely carry-over effect of SCP. Finally, we measured 50-kHz and 22-kHz ultrasonic vocalizations (USVs) as reliable markers of positive and negative emotionality, respectively. We found that SCP increased locomotion during OF1 and reduced rearing on OF1-OF4, causing no disruption in habituation over tests. SCP prevented the increase of total grooming time by inhibiting complex grooming subtypes and promoting short cephalic sequences. Despite the SCP-induced alterations on grooming agreed with our hypotheses, those changes may have resulted from a motor impairment that could have also affected rearing behavior. Additionally, SCP suppressed 50-kHz USVs while marginally increased 22-kHz calls. Once SCP was withdrawn, rearing, grooming, and some 50-kHz USVs subtypes returned to VEH levels, suggesting that novelty habituation occurred despite the SCP administration. Altogether, that mixed profile of SCP-induced behavioral changes may derive from the complex interplay between the contrasting action of SCP on different brain regions and the doses here used., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Cholinergic neuromodulation is known to play a key role in visual working memory (VWM) - keeping relevant stimulus representations available for cognitive processes for short time periods (up to a few minutes). Despite the growing body of evidence on how the neural and cognitive mechanisms of VWM dynamically change over retention time, there is mixed evidence available on cholinergic effects as a function of VWM delay period in non-human primates. Using the delayed matching to sample VWM task in rhesus macaques (N = 6), we aimed to characterize VWM maintenance in terms of performance changes as a function of delay duration (across a wide range of delays from 1 to 76 s). Then, we studied how cholinergic neuromodulation influences VWM maintenance using the muscarinic receptor antagonist scopolamine administered alone as transient amnestic treatment, and in combination with two doses of the acetylcholinesterase inhibitor donepezil, a widely used Alzheimer's medication probing for the reversal of scopolamine-induced impairments. Results indicate that scopolamine-induced impairments of VWM maintenance are delay-dependent and specifically affect the 15-33 s time range, suggesting that scopolamine worsens the normal decay of VWM with the passage of time. Donepezil partially rescued the observed scopolamine-induced impairments of VWM performance. These results provide strong behavioral evidence for the role of increased cholinergic tone and muscarinic neuromodulation in the maintenance of VWM beyond a few seconds, in line with our current knowledge on the role of muscarinic acetylcholine receptors in sustained neural activity during VWM delay periods., (Copyright © 2020 The Author(s). Published by Elsevier B.V. All rights reserved.)

The rostromedial tegmental nucleus (RMTg) receives inputs from the laterodorsal tegmental and pedunculopontine tegmental nuclei, the two principle brainstem cholinergic nuclei. We tested the effects of RMTg M3 and M4 muscarinic cholinergic receptor antagonism in a conditioned place preference (CPP) paradigm in mice. RMTg infusions of the M3 muscarinic cholinergic receptor antagonist 1,1-Dimethyl-4-diphenylacetoxypiperidinium iodide (4-DAMP) do not result in the acquisition of CPP but increase locomotor activation. By contrast, RMTg infusions of the M4 muscarinic cholinergic receptor antagonist Tropicamide result in the acquisition of CPP but do not increase locomotor activation. The rewarding effects of RMTg Tropicamide infusions are dopamine-dependent as systemic pre-treatment with the broad-spectrum dopamine receptor antagonist flupenthixol prevents the acquisition of CPP induced by RMTg Tropicamide infusions. Under conditions of systemic dopamine receptor blockade, RMTg Tropicamide infusions significantly increase locomotor activation. These data provide further support for an important role of endogenous cholinergic input to the RMTg in reward function and suggest that the contributions of RMTg cholinergic input to rewarding and locomotor-activating effects involve differential contributions of RMTg M4 and M3 muscarinic receptors, respectively., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Filial imprinting in precocial birds is a useful model for studying memory formation in early learning. The intermediate medial mesopallium (IMM) in the dorsal telencephalon is one of the critical brain regions where the releases of several neurotransmitters increase after the start of imprinting training. Among the increased neurotransmitters, the role of acetylcholine in imprinting has remained unclear. Acetylcholine in the mammalian brain plays an important role in encoding new memories. The muscarinic acetylcholine receptor subtype 1 (M 1 receptor) and subtype 3 (M 3 receptor) in the hippocampus and cortex of mammalian brain have been shown to be necessary for memory encoding. In this study, we examined whether the imprinting acquisition in chick can be impaired by injecting muscarinic acetylcholine receptor (mAChR) antagonist scopolamine into the bilateral IMM. We show that the injection of scopolamine decreased the preference for the imprinting object in the test, but did not affect the number of approaches to the imprinting object during training. Immunoblotting and immunohistochemistry revealed that M 3 receptors were expressed in the IMM. Our data suggest that acetylcholine is involved in the memory formation of imprinting through M 3 receptors in the IMM. The scopolamine-injected chicks may be useful as an animal model for dementia such as Alzheimer's disease., (Copyright © 2019 Elsevier B.V. All rights reserved.)

The purpose of the work was to study the role of anti-inflammatory cytokines in memory processing in a healthy brain. Wistar rats were trained to perform a task with positive (food) reinforcement; and then the task performance was tested after intraventricular injection of IL-10 or TGF-β1. A microinjection into the brain of either of the two cytokines did not affect the performance of the task and did not have an anti-amnesic effect when the retrieval was deteriorated with scopolamine. In addition, endogenous levels of IL-10 and TGF-β1 were determine in the prefrontal cortex and in the hippocampus after one and two training sessions, consisting of 10 runs each. The level of IL-10 did not change after training both in the prefrontal cortex and in the hippocampus. Endogenous level of TGF-β1 increased in the neocortex after the first training session, the second session, and recovered to the normal level three days after training. In contrast, in the hippocampus, the level of TGF-β1 was decreased: maximally after the first training session in the right hippocampus and after the second training session in the left one. Given the role of the prefrontal cortex in memory processing, we assume that a specific increase of TGF-β1 in the prefrontal cortex may indicate involvement in memory trace consolidation., (Copyright © 2019. Published by Elsevier B.V.)

Treatment of fasted mice and rats with the nonselective muscarinic antagonist, scopolamine or atropine, causes convulsions after food intake. This study evaluated the effect of fasting on the expression of M 1 and M 2 muscarinic receptors in the brain regions, the relationship between receptor expression and seizure stages, and the muscarinic receptor subtype which plays a role in the occurrence of convulsions. Mice were grouped as allowed to eat ad lib (fed) and deprived of food for 24h (fasted). Fasted animals developed convulsions after being treated with scopolamine (60%) or the selective M 1 receptor antagonist pirenzepine (10mg/kg; 20% and 60mg/kg; 70%) and given food. Fasting increased expression of M 1 receptors in the frontal cortex and M 2 receptors in the hippocampus, but produced no change in the expression of both receptors in the amygdaloid complex. Food intake after fasting decreased M 1 receptor expression in the frontal cortex and M 1 and M 2 receptor expression in the hippocampus. Seizure severity was uncorrelated with muscarinic receptor expression in the brain regions. Taken together, these findings provide evidence for the role of M 1 muscarinic receptor antagonism and fasting-induced increases in M 1 and M 2 expression possible underlying mechanism in the occurrence of convulsions in fasted animals., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Repeated administration of morphine profoundly influences the dopaminergic and cholinergic systems in the nucleus accumbens [including the shell of the nucleus accumbens (NAcS)]. Further, dopamine release is regulated by the cholinergic system, especially the M 4 receptor. Drug priming is one of the main factors that induces relapse in drug addiction. The present study first investigated how activation of the M 4 receptor in the NAcS affects the expression of morphine-induced behavioral sensitization, through the administration of an M 4 agonist (LY2033298) and antagonist (tropicamide), as well as a combination of an acetylcholinesterase inhibitor and M 4 antagonist (huperzine-A + tropicamide). Additionally, the influence of a dopamine receptor agonist, in conjunction with an M 4 agonist (i.e., SKF38393 + LY2033298), was also examined. Behavioral sensitization was established by exposure to 5 mg/kg morphine once every three days for a total of three exposures. The expression of behavioral sensitization was challenged by 5 mg/kg morphine. Results showed that (1) microinjection of the M 4 receptor agonist LY2033298 (0.2 μg/side), but not the antagonist tropicamide (5, 10, or 20 μM/side) into the NAcS blocked the expression of behavioral sensitization; (2) tropicamide (20 μM/side) reversed the inhibition effect of huperzine-A on this behavior; and (3) SKF38393 (1 μg/side) reversed the inhibitory effect of LY2033298 on the expression of morphine-induced behavioral sensitization. These results suggest that the cholinergic M 4 receptor in the NAcS plays an important role in the morphine-induced expression of behavioral sensitization through the regulation of dopamine function in rats., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Mesopontine and basal forebrain cholinergic neurons are involved in the control of behavioral states and cognitive functions. Animals treated with cholinergic muscarinic receptor antagonists display a dissociated state characterized by behavioral wakefulness (W) associated with high amplitude slow oscillations and spindles in the electroencephalogram (EEG), similar to those that occur during non-REM (NREM) sleep. Oscillations in the gamma frequency band (≈ 40 Hz) of the EEG also play a critical role during W and cognition. Hence, the present study was conducted to determine the effect of muscarinic antagonists on the EEG gamma band power and coherence. Five cats were implanted with electrodes in different cortices to monitor the EEG. The effects of atropine and scopolamine on power and coherence within the low gamma frequency band (30-45 Hz) from pairs of EEG recordings were analyzed and compared to gamma activity during sleep and W. Muscarinic antagonists induced a NREM sleep-like EEG profile that was accompanied by a large increase in gamma power and coherence. The values of gamma coherence were similar to that occurring during alert W (AW), and greater than in quiet W, NREM and REM sleep. We conclude that under atropine or scopolamine, functional interactions between cortical areas in the gamma frequency band remain high, as they are during AW. This significant functional connectivity at high frequency may explain why the animals remain awake in spite of the presence of slow waves and spindles., (Copyright © 2018 Elsevier B.V. All rights reserved.)

The capacity to recognize objects from different view-points or angles, referred to as view-invariance, is an essential process that humans engage in daily. Currently, the ability to investigate the neurobiological underpinnings of this phenomenon is limited, as few ethologically valid view-invariant object recognition tasks exist for rodents. Here, we report two complementary, novel view-invariant object recognition tasks in which rodents physically interact with three-dimensional objects. Prior to experimentation, rats and mice were given extensive experience with a set of 'pre-exposure' objects. In a variant of the spontaneous object recognition task, novelty preference for pre-exposed or new objects was assessed at various angles of rotation (45°, 90° or 180°); unlike control rodents, for whom the objects were novel, rats and mice tested with pre-exposed objects did not discriminate between rotated and un-rotated objects in the choice phase, indicating substantial view-invariant object recognition. Secondly, using automated operant touchscreen chambers, rats were tested on pre-exposed or novel objects in a pairwise discrimination task, where the rewarded stimulus (S+) was rotated (180°) once rats had reached acquisition criterion; rats tested with pre-exposed objects re-acquired the pairwise discrimination following S+ rotation more effectively than those tested with new objects. Systemic scopolamine impaired performance on both tasks, suggesting involvement of acetylcholine at muscarinic receptors in view-invariant object processing. These tasks present novel means of studying the behavioral and neural bases of view-invariant object recognition in rodents., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Angiotensin II (ANG II) is a typical facilitatory stimulus for sodium appetite. Surprisingly, hyperosmolarity and central cholinergic stimulation, two classical antinatriorexigenic stimuli, also facilitate NaCl intake when they are combined with injections of the α 2 -adrenoceptor/imidazoline agonist moxonidine into the lateral parabrachial nucleus (LPBN). In the present study, we tested the relative importance of central angiotensinergic and cholinergic mechanisms for the control of water and NaCl intake by combining different dipsogenic or natriorexigenic stimuli with moxonidine injection into the LPBN. Adult male Holtzman rats (n=9-10/group) with stainless steel cannulas implanted in the lateral ventricle and LPBN were used. Bilateral injections of moxonidine (0.5 nmol) into the LPBN increased water and 0.3M NaCl intake in rats that received furosemide+captopril injected subcutaneously, ANG II (50ng) or carbachol (cholinergic agonist, 4 nmol) injected intracerebroventricularly (icv) or 2M NaCl infused intragastrically (2ml/rat). Losartan (AT 1 antagonist, 100μg) or atropine (muscarinic antagonist, 20 nmol) injected icv abolished the effects on water and 0.3M NaCl of moxonidine combined to either 2M NaCl intragastrically or carbachol icv. However, atropine icv did not change 0.3M NaCl intake produced by direct central action of ANG II like that induced by ANG II icv or furosemide+captopril combined with moxonidine into the LPBN. The results suggest that different stimuli, including hyperosmolarity and central cholinergic stimulation, share central angiotensinergic activation as a common mechanism to facilitate sodium intake, particularly when they are combined with deactivation of the LPBN inhibitory mechanisms., (Copyright © 2017 Elsevier B.V. All rights reserved.)

The phenomenon of utilizing information acquired in the past to make decision and performance in present depends on memory retrieval, which is affected in retrograde amnesia. Role of cholinergic receptors in memory retrieval is not much explored. In this study we evaluated the gender specific role of cholinergic receptors, i.e. muscarinic and nicotinic receptors, in memory retrieval in young Balb/c mice. Acute (only one injection, 30min before test) and sub-chronic (five days) muscarinic blockade (using scopolamine=1mg/kg) before test impaired retrieval of contextual fear memory in male (31.45±5.39% and 33.36±3.78% respectively) and female mice (22.88±5.73%; P<0.05), except sub-chronically treated female group (33.31±4.90%; P>0.05). Only sub-chronic nicotinic receptor antagonism (using methyllycaconitine MLA=87.5μg/kg and dihydro β erythroidine DHβE=1mg/kg) in female showed significantly higher freezing response than control during contextual fear memory retrieval (60.85±7.71% and 40.91±7.53% respectively; P<0.001). Acute and sub-chronic muscarinic antagonism (but not nicotinic antagonism) impaired spatial memory retrieval in male (P<0.05) but not in female mice (P>0.05). There was no effect of acute and sub-chronic cholinergic receptor antagonism on discriminating novel object from the familiar one in male and female mice, however, nicotinic receptor blockade affected the working memory of all male and female mice on test day compared to the training sessions. Our results suggested that cholinergic receptors involvement in retrieving spatial and fear memories depends on the age of the memory and gender., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Muscarinic M 1 /M 4 receptor stimulation can reduce abuse-related effects of cocaine and may represent avenues for treating cocaine addiction. Muscarinic antagonists can mimic and enhance effects of cocaine, including discriminative stimulus (S D ) effects, but the receptor subtypes mediating those effects are not known. A better understanding of the complex cocaine/muscarinic interactions is needed to evaluate and develop potential muscarinic-based medications. Here, knockout mice lacking M 1 , M 2 , or M 4 receptors (M 1 -/- , M 2 -/- , M 4 -/- ), as well as control wild-type mice and outbred Swiss-Webster mice, were trained to discriminate 10mg/kg cocaine from saline. Muscarinic receptor antagonists with no subtype selectivity (scopolamine), or preferential affinity at the M 1 , M 2 , or M 4 subtype (telenzepine, trihexyphenidyl; methoctramine, AQ-RA 741; tropicamide) were tested alone and in combination with cocaine. In intact animals, antagonists with high affinity at M 1 /M 4 receptors partially substituted for cocaine and increased the S D effect of cocaine, while M 2 -preferring antagonists did not substitute, and reduced the S D effect of cocaine. The cocaine-like effects of scopolamine were absent in M 1 -/- mice. The cocaine S D attenuating effects of methoctramine were absent in M 2 -/- mice and almost absent in M 1 -/- mice. The findings indicate that the cocaine-like S D effects of muscarinic antagonists are primarily mediated through M 1 receptors, with a minor contribution of M 4 receptors. The data also support our previous findings that stimulation of M 1 receptors and M 4 receptors can each attenuate the S D effect of cocaine, and show that this can also be achieved by blocking M 2 autoreceptors, likely via increased acetylcholine release., (Copyright © 2017 Elsevier B.V. All rights reserved.)

In the present study we investigated whether stimulation of muscarinic acetylcholine (mACh) receptors in the ventral tegmental area (VTA) plays a role in the acquisition of food-based conditioned approach learning. Rats were exposed to 3 (in Experiment 1) or 7 (in Experiment 2) conditioning sessions in which 30, randomly presented light (CS) presentations were paired with delivery of food pellets (US), followed by one session with no light or food and finally one CS-only test session with only light stimulus presentations. Bilateral microinjections of scopolamine (a mACh receptor antagonist) were made either prior to each conditioning session (Experiment 1; to test effects on acquisition) or prior to the CS-only test (Experiment 2; to test effects on performance of the learned response). Scopolamine produced a dose-related significant reduction in the acquisition of conditioned approach but had no effect on its performance. These results suggest that mACh receptor stimulation in the VTA plays a necessary role in the acquisition of reward-related learning., (Copyright © 2017 Elsevier B.V. All rights reserved.)

The mechanisms of post-stroke neurogenesis in the subventricular zone (SVZ) are unclear. However, neural stem cell-intrinsic and neurogenic niche mechanisms, as well as neurotransmitters, have been shown to play important roles in SVZ neurogenesis. Recently, a previously unknown population of choline acetyltransferase (ChAT) + neurons residing in rodent SVZ were identified to have direct control over neural stem cell proliferation by indirectly activating fibroblast growth factor receptor (FGFR). This finding revealed possible neuronal control over SVZ neurogenesis. In this study, we assessed whether these ChAT + neurons also participate in stroke-induced neurogenesis. We used a permanent middle cerebral artery occlusion (MCAO) model produced by transcranial electrocoagulation in mice, atropine (muscarinic cholinergic receptor [mAchR] antagonist), and donepezil (acetylcholinesterase inhibitor) to investigate the role of ChAT + neurons in stroke-induced neurogenesis. We found that mAchRs, phosphorylated protein kinase C (p-PKC), and p-38 levels in the SVZ were upregulated in mice on day 7 after MCAO. MCAO also significantly increased the number of BrdU/doublecortin-positive cells and protein levels of phosphorylated-neural cell adhesion molecule and mammalian achaete scute homolog-1. FGFR was activated in the SVZ, and doublecortin-positive cells increased in the peri-infarction region. These post-stroke neurogenic effects were enhanced by donepezil and partially decreased by atropine. Neither atropine nor donepezil affected peri-infarct microglial activation or serum concentrations of TNF-α, IFN-γ, or TGF-β on day 7 after MCAO. We conclude that ChAT + neurons in the SVZ may participate in stroke-induced neurogenesis, suggesting a new mechanism for neurogenesis after stroke., (Copyright © 2016 Elsevier B.V. All rights reserved.)

To determine whether 3-furan-2-yl-N-p-tolyl-acrylamide (PAM-2), a positive allosteric modulator of α7 nicotinic acetylcholine receptors (nAChRs), improves memory processes, passive avoidance tests were conducted in male mice after acute and chronic treatments. To determine the neuronal mechanisms underlying the promnesic activity elicited by PAM-2, the effect of this ligand on α7 nAChR up-regulation and ERK1/2 phosphorylation was assessed in the hippocampus and prefrontal cortex. The results indicate that: (1) PAM-2 improves memory acquisition/consolidation after acute treatment (Day 2) and memory consolidation after chronic treatment (Day 22). Although no effect was observed on α7 nAChR up-regulation, the chronic, but not acute, PAM-2 treatment increases ERK1/2 kinase phosphorylation, (2) the promnesic activity of PAM-2 was inhibited by methyllycaconitine, a selective α7-antagonist, confirming the role of α7 nAChRs, (3) a synergistic (acute) effect was observed between inactive doses of PAM-2 (0.1 mg/kg) and DMXBA (0.3 mg/kg), a selective α7-agonist, and (4) PAM-2 reversed the memory impairment elicited by scopolamine, a muscarinic antagonist. The results demonstrate that PAM-2 presents promnesic activity mediated by α7 nAChRs, and is able to trigger ERK1/2 phosphorylation only after chronic treatment., (Copyright © 2016 Elsevier B.V. All rights reserved.)

The mesolimbic dopamine (DA) system is known to play a role in cue-mediated reward-seeking for natural rewards and drugs of abuse. Specifically, cholinergic and glutamatergic receptors in the ventral tegmental area (VTA) have been shown to regulate cue-induced drug-seeking. However, the potential role of these VTA receptors in regulating cue-induced reward seeking for natural rewards is unknown. Here, we examined whether blockade of VTA acetylcholine receptors (AChRs) and N-methyl-d-aspartate receptors (NMDARs) would alter cue-induced sucrose seeking in male Sprague-Dawley rats. Subjects underwent 10 days of sucrose self-administration training (fixed ratio 1 schedule) followed by 7 days of forced abstinence. On withdrawal day 7, rats received bilateral VTA infusion of vehicle, the muscarinic AChR antagonist scopolamine (2.4 or 24 μg/side), the nicotinic AChR antagonist mecamylamine (3 or 30 μg/side), or the NMDAR antagonist AP-5 (0.1 or 1 μg/side) immediately prior to examination of cue-induced sucrose-seeking. Scopolamine infusion led to robust attenuation, but did not completely block, sucrose-seeking behavior. In contrast, VTA administration of mecamylamine or AP-5 did not alter cue-induced sucrose-seeking. Together, the data suggest that VTA muscarinic AChRs, but not nicotinic AChRs nor NMDARs, facilitate the ability of food-associated cues to drive seeking behavior for a food reward., (Copyright © 2015 Elsevier B.V. All rights reserved.)

GPR37, also known as parkin associated endothelin-like receptor (Pael-R), is an orphan GPCR that aggregates intracellularly in a juvenile form of Parkinson's disease. However, little is known about the function of this orphan receptor. Here, using a model for parkisonian tremor, the pilocarpine-induced tremulous jaw movements (TJMs), we show that the deletion of GPR37 attenuated the TJMs in response to this cholinomimetic. Interestingly, the control that adenosine A2A receptor exerted over TJMs was lost in the absence of GPR37, thus pointing to a pivotal role of this orphan receptor in the adenosinergic control of parkinsonian tremor., (Copyright © 2015 Elsevier B.V. All rights reserved.)

The action of PACAP-38 was studied by measuring the anxiogenic-anxiolytic behavior of rats in an elevated plus maze. PACAP-38 was administered into the lateral brain ventricle and the behavior of the animals was measured 3h later. The possible involvement of transmitters was measured by pretreating the animals with receptor blockers which alone did not influence the task, but in the doses used were effective with other neuropeptides. The receptor antagonist PACAP 6-38 (a PAC 1/VPAC2 receptor antagonist of PACAP-38 receptor), haloperidol (a non-selective dopamine receptor antagonist), phenoxybenzamine (an α1/α2β-adrenergic receptor antagonist), propranolol(a β-adrenergic receptor antagonist), bicuculline (a gamma-aminobutyric acid subunit A receptor antagonist), methysergide (a nonselective 5-HT2 serotonergic receptor antagonist), atropine (a nonselective muscarinic acetylcholine receptor antagonist), naloxone (a nonselective opioid receptor antagonist) and nitro-l-arginine which acts by blocking the enzyme nitric oxide synthase, thereby blocking the nitric oxide synthesis, were tested. The following parameters were measured: the time spent in open arms/the time spent in total entries. PACAP-38 decreased the ratio of time spent in open arms to the time spent in total entries, indicating anxiogenic action. The total number of entries was not altered significantly either by PACAP-38 or by the receptor blockers. The following receptor blockers diminished the action of PACAP-38: PACAP 6-38,haloperidol, methysergide, naloxone and nitro-l-arginine. Pretreatment with atropine, phenoxybenzamine, propranolol and bicuculline did not influence the action of PACAP-38 on the time spent in open arms. The results demonstrate that PACAP-38 administered into the lateral brain ventricle exerted anxiogenic action at 3 h following treatment. Pretreatment of the animals with various receptor blockers indicated that a nonselective dopaminergic receptor antagonist, 5HT2 serotonergic and opioid receptors, nitric oxide and PAC1 receptors are involved in the anxiogenic action induced by PACAP-38., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Disorders characterized by disturbed cholinergic signaling, such as schizophrenia, exhibit impaired performance on measures of real-world cost/benefit decision-making. Whether the cholinergic system contributes to the choice deficits observed is currently unknown. We therefore determined the effects of broad-acting agonists and antagonists at the nicotinic and muscarinic receptor on decision making, as measured by the rodent gambling task (rGT). Given the anatomical and functional connectivity of the cholinergic and dopaminergic systems, we also sought to modulate amphetamine's previously reported effect on rGT performance via the cholinergic system. Male rats were trained on the rGT, during which animals chose from four different options. The optimal strategy on the rGT is to favor options associated with smaller immediate rewards and less punishment/loss. Impulsive action was also measured by recording the number of premature responses made. Performance on the rGT was assessed following acute treatment with the muscarinic receptor agonist oxotremorine, the muscarinic receptor antagonist scopolamine, nicotine, and the nicotinic receptor antagonist mecamylamine. Similar to the effect produced by amphetamine, muscarinic receptor antagonism with scopolamine (0.1mg/kg) impaired decision making, albeit to a lesser degree. Prior muscarinic agonism with oxotremorine was unable to attenuate amphetamine's effects on rGT performance. Oxotremorine, nicotine, and mecamylamine did not affect the choice profile. We therefore conclude that modulation of the muscarinic, but not nicotinic, receptor system can affect decision making under conditions of risk and uncertainty. Such findings contribute to a broader understanding of the cognitive deficits observed in disorders in which cholinergic signaling is compromised., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Urocortin 3 (Ucn 3) was found to impair passive avoidance learning in male and female mice. The possible involvement of transmitters in the action, the animals were pretreated with the following receptor antagonists in doses which alone could not influence the measurement. Haloperidol (a D2, dopamine receptor antagonist), phenoxybenzamine (a nonselective α1-adrenergic receptor antagonist), atropine (a nonselective muscarinic acetylcholine receptor antagonist), bicuculline (a γ-aminobutyric acid subunit A receptor antagonist), nitro-L-arginine (a nitric oxide synthase inhibitor), antalarmin (a CRF1 receptor antagonist) and astressin 2B (a CRF2 receptor antagonist). Haloperidol, phenoxybenzamine, bicuculline, atropine, nitro-L-arginine and astressin 2B prevented the action of Ucn 3, in both sexes, whereas antalarmin exerted no action in either male or female animals., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Anxiety is one of the most prevalent neuropsychiatric disorders, and little is known about its pathogenesis. In order to investigate the neural mechanisms of this mental disorder, we used rat behavior in the elevated plus-maze as an animal model of anxiety and the insular cortex (insula) as a brain target. The microinjection of non-selective and selective M1 and M4 muscarinic acetylcholine receptor (mAChR) agonists or antagonists was used to explore whether the insular muscarinic receptor and its subtypes regulate levels of anxiety. The results showed that both non-selective and selective M1 and M4 mAChR agonists increased the time spent on exploring in the open arms, whereas antagonists decreased exploration. Our results indicate that activation of insular mAChRs could produce anxiolytic effects, whereas inhibition of insular mAChRs could increase anxiety. We concluded that the insular muscarinic system plays a role in the modulation of anxiety, and dysfunction of mAChR signaling may be involved in the mechanism of anxiogenesis., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The influence of systemic administration of cholinesterase inhibitors, donepezil and rivastigmine on the acquisition, expression, and reinstatement of morphine-induced conditioned place preference (CPP) was examined in rats. Additionally, this study aimed to compare the effects of donepezil, which selectively inhibits acetylcholinesterase, and rivastigmine, which inhibits both acetylcholinesterase and butyrylcholinesterase on morphine reward. Morphine-induced CPP (unbiased method) was induced by four injections of morphine (5 mg/kg, i.p.). Donepezil (0.5, 1, and 3 mg/kg, i.p.) or rivastigmine (0.03, 0.5, and 1 mg/kg, i.p.) were given 20 min before morphine during conditioning phase and 20 min before the expression or reinstatement of morphine-induced CPP. Our results indicated that both inhibitors of cholinesterase attenuated the acquisition and expression of morphine CPP. The results were more significant after rivastigmine due to a broader inhibitory spectrum of this drug. Moreover, donepezil (1 mg/kg) and rivastigmine (0.5 mg/kg) attenuated the morphine CPP reinstated by priming injection of 5mg/kg morphine. These properties of both cholinesterase inhibitors were reversed by mecamylamine (3 mg/kg, i.p.), a nicotinic acetylcholine receptor antagonist but not scopolamine (0.5 mg/kg, i.p.), a muscarinic acetylcholine receptor antagonist. All effects of cholinesterase inhibitors were observed at the doses that had no effects on locomotor activity of animals. Our results suggest beneficial role of cholinesterase inhibitors in reduction of morphine reward and morphine-induced seeking behavior. Finally, we found that the efficacy of cholinesterase inhibitors in attenuating reinstatement of morphine CPP provoked by priming injection may be due to stimulation of nicotinic acetylcholine receptors., (Copyright © 2014 Elsevier B.V. All rights reserved.)

The widespread distribution of apelin-13 and apelin receptors in the brain and periphery suggests an important function of this neuropeptide in regulatory processes in the organism. In previous work we found that apelin-13 facilitates the consolidation of passive avoidance learning in rats. In the present work we demonstrate that apelin-13 exerts anxiolytic action in an elevated plus maze in mice. In order to assess the possible involvement of transmitters in this action, the animals were pretreated with the following receptor blockers in doses which themselves did not influence the behavioral paradigm: atropine (a nonselective muscarinic acetylcholine receptor antagonist), haloperidol (a D2, D3, D4 dopamine receptor antagonist), phenoxybenzamine (a nonselective α1-adrenergic receptor antagonist), methysergide (a nonselective 5-HT2 serotonergic receptor antagonist), propranolol (a β-adrenergic receptor antagonist), naloxone (a nonselective opioid receptor antagonist) and bicuculline (a γ-aminobutyric acid subunit A receptor antagonist. Phenoxybenzamine, haloperidol, propranolol and methysergide prevented the action of apelin-13, whereas atropine, naloxone and bicuculline were ineffective. The data suggest that apelin-13 elicits its anxiolytic action via α-adrenergic, dopaminergic, β-adrenergic and 5-HT2 serotonergic mediation., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Neuromedin U (NmU) is a widely distributed and multifunctional peptide in the central nervous system and the peripheral tissues. Little is know about the mechanisms of NmU on brain functions. The rodent isoform of the NmU, NmU-23, has been shown to have anxiolytic effects involved in the β-adrenergic and cholinergic nervous systems in elevated plus maze test. NmU-23 was tested for antidepressant-like effects in modified forced swimming test (FST) in mice and furthermore, the involvement of the adrenergic, serotonergic, cholinergic, dopaminergic or gaba-ergic receptors in the antidepressant-like effect of NmU-23 was studied in modified mice FST. Mice were pretreated with a non-selective α-adrenergic receptor antagonist phenoxybenzamine, an α1/α2β-adrenergic receptor antagonist, prazosin, an α2-adrenergic receptor antagonist, yohimbine, a β-adrenergic receptor antagonist, propranolol, a mixed 5-HT1/5-HT2 serotonergic receptor antagonist, methysergide, a non-selective 5-HT2 serotonergic receptor antagonist, cyproheptadine, nonselective muscarinic acetylcholine receptor antagonist, atropine, D2,D3,D4 dopamine receptor antagonist, haloperidol or γ-aminobutyric acid subunit A (GABAA) receptor antagonist, bicuculline. NmU-23 showed the antidepressant-like effects by decreasing the immobility time and increasing the climbing and swimming time. Prazosin, haloperidol, and bicuculline prevented the effects of NmU-23 on the climbing and swimming time. Methysergide and cyproheptadine prevented the effects of NmU-23 on the immobility, swimming and climbing time. Atropine prevented the effects of NmU-23 on the climbing time. Phenoxybenzamine, yohimbine and propranolol did not change the effects of NmU-23. The results demonstrated that the antidepressant-like effect of NmU-23 is mediated, at least in part, by an interaction of the α2-adrenergic, 5-HT1-2 serotonergic, D2,D3,D4 dopamine receptor, muscarinic acetylcholine receptors and γ-aminobutyric acid subunit A (GABAA) receptor in a modified mouse FST., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The entorhinal cortex receives inputs from sensory and associational cortices, as well as a substantial input from midbrain dopaminergic neurons. Dopamine is likely to modulate the responsiveness of entorhinal cortex neurons to sensory inputs, and excitatory synaptic responses in layers I/II of the entorhinal cortex in vitro can be either facilitated or suppressed by dopamine depending upon the concentration applied. Rewarding stimulation of the lateral hypothalamus leads to activation of dopamine neurons, and the present study evaluated the effect of rewarding stimulation on synaptic responses in the lateral entorhinal cortex evoked by stimulation of the primary olfactory (piriform) cortex in behaving rats. Rewarding brain stimulation reduced the amplitude of synaptic responses in the entorhinal cortex evoked by single pulses delivered to the piriform cortex at intervals of 100-500 ms following the train. Synaptic responses were suppressed when stimulation trains were delivered at a fixed interval, or when trains were initiated by the animal pressing a bar. The suppression depended on the strength of stimulation trains; delivery of higher frequency trains that were sufficient to induce maximal, or 50% of maximal, rates of bar-pressing resulted in significant suppression effects, but lower frequency trains did not. Systemic administration of the dopamine D2 receptor antagonist eticlopride, but not the D1 receptor antagonist SCH23390 or the muscarinic antagonist scopolamine, blocked the suppression of synaptic responses. Results suggest that rewarding brain stimulation leads to a phasic increase in dopamine in the entorhinal cortex resulting in a D2 receptor-dependent suppression of excitatory synaptic responses, and that a similar synaptic modulation may be induced by stimuli associated with appetitive motivation and reward., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Behavioral sensitization to the repeated administration of a psychostimulant presumably plays a key role in the pathogenesis of addiction and schizophrenia. Among other psychostimulants, 3,4-methylenedioxymethamphetamine (MDMA) is known to produce behavioral sensitization, too, but its mechanism of action is still not fully understood. Along with the strong release of catecholamines and serotonin, MDMA exerts actions at additional transmitter systems, including acetylcholine (ACh). To identify the cholinergic involvement in the development and expression of MDMA-induced sensitization, rats were treated daily with MDMA (5.0 mg/kg), MDMA plus the muscarinic antagonist atropine (4.28 mg/kg), or MDMA plus the nicotinic antagonist mecamylamine (1.0 mg/kg) for 13 consecutive days. The results show that atropine co-treatment was able to block the development of behavioral sensitization to MDMA, measured as horizontal activity and rearing, whereas mecamylamine did not. Pharmacological challenge with MDMA alone increased the locomotion in all substance pretreated groups with the MDMA plus atropine group showing the lowest values. The second challenge with MDMA plus atropine showed a decrease in locomotor behavior in the MDMA- and an increase in the MDMA plus atropine pretreated groups, resulting in similar levels of activity for both groups. A control experiment revealed no change in horizontal activity and rearing when only the cholinergic antagonists (atropine; mecamylamine) were administered. This is the first study that shows a substantial role of muscarinic receptors for the development of behavioral sensitization to MDMA., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Inverse agonism at the benzodiazepine site of α(5) subunit-containing GABA(A) receptors is an attractive approach for the development of putative cognition-enhancing compounds, which are still far from clinical application. Several ligands with binding and/or functional selectivity for α(5) GABA(A) receptors have been synthesized and tested in a few animal models. PWZ-029 is an α(5) GABA(A) selective inverse agonist whose memory enhancing effects were demonstrated in the passive avoidance task in rats and in Pavlovian fear conditioning in mice. In the present study we investigated the effects of PWZ-029 administration in novel object recognition test and Morris water maze, in normal and scopolamine-treated rats. All the three doses of PWZ-029 (2, 5 and 10 mg/kg) improved object recognition after the 24-h delay period, as shown by significant differences between the exploration times of the novel and old object, and the respective discrimination indices. PWZ-029 (2 mg/kg) also successfully reversed the 0.3 mg/kg scopolamine-induced deficit in recognition memory after the 1-h delay. In the Morris water maze test, PWZ-029 (5, 10 and 15 mg/kg) did not significantly influence swim patterns, either during five acquisition days or during the treatment-free probe trial. PWZ-029 (2, 5 and 10 mg/kg) also proved to be ineffective in the reversal of the 1mg/kg scopolamine-induced memory impairment in the water maze. The present mixed results encourage use of a variety of tests and experimental conditions in order to increase the predictability of preclinical testing of selective α(5) GABA(A) inverse agonists., (Copyright © 2012 Elsevier B.V. All rights reserved.)

We designed this study to investigate effects of intra-nucleus accumbens (intra-NAc) infusions of GABA(A) receptors agonist (muscimol) and antagonist (bicuculline) by themselves and their interaction with scopolamine on inhibitory avoidance (IA) memory performance. This study used a step-through IA task to assess memory in male Wistar rats. The results showed that post-training intra-NAc infusions of muscimol at doses of 0.01 and 0.02 μg/rat significantly impaired IA memory performance, while bicuculline (0.1, 0.2 and 0.4 μg/rat) had no effect. Post-training intra-NAc infusions of scopolamine at dose of 0.5 μg/rat impaired IA memory performance by itself, and in combination with an ineffective dose of muscimol increased impairment of IA memory performance. The results also showed that post-training intra-NAc infusions of bicuculline prevented the impairing effect of higher doses of scopolamine on IA memory performance. Pre-test intra-NAc infusions of muscimol (0.01 and 0.02 μg/rat) and bicuculline (0.001, 0.01 and 0.1 μg/rat) impaired IA memory performance. The results also revealed that pre-test intra-NAc infusions of scopolamine (0.25 and 0.5 μg/rat) impaired IA memory performance, and its co-infusions with an ineffective dose of muscimol (0.005 μg/rat) increased impairment of IA memory performance. Interestingly, pre-test intra-NAc infusions of bicuculline impaired performance by itself, but did not prevent the impairing effect of scopolamine. It can be concluded that GABA(A) and muscarinic receptors of the NAc may have an interaction in modulation of IA memory performance, which may result from affecting output neurons in the NAc directly or indirectly via cholinergic and GABAergic interneurons., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Central cholinergic transmission has long been implicated in various cognitive processes, including memory acquisition, consolidation, and attentional processes. Here, we examined the role of muscarinic receptors in visual discrimination performance under conditions of altered visual information availability. Adult rats were trained to discriminate two visual cues (indicating the presence and absence of a hidden escape platform, respectively) in a water maze-based, trapezoidal-shaped apparatus. Following task acquisition, testing continued with two types of trials: regular trials (RTs; both visual cues present, identical to training conditions) and probe trials (PTs; only one of the two cues present). In Experiment 1, removal of one visual cue on PTs impaired discrimination performance. Moreover, scopolamine administration (0.125-1.0 mg/kg, i.p.) tended to further suppress performance in a dose-dependent manner on PTs, while discriminations on RTs were left intact. In Experiment 2, these results were confirmed and extended by showing that PT (one visual cue) performance could improve with training in undrugged, but not in scopolamine-treated rats. Together, these experiments reveal that visual discrimination performance of rats benefits from the concurrent availability of two visual cues that provide complimentary and consistent information. Furthermore, muscarinic receptors are particularly important under conditions of reduced visual information availability, as well as in the adoption of new behavioral strategies, such as switching from two-cue to single-cue guided navigation., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Evidences indicate that inhibition of central Renin angiotensin system (RAS) ameliorates memory impairment in animals and humans. Earlier we have reported involvement of central angiotensin converting enzyme (ACE) in streptozotocin induced neurodegeneration and memory impairment. The present study investigated the role of central ACE in cholinergic neurotransmission, brain energy metabolism and cerebral blood flow (CBF) in model of memory impairment induced by injection of scopolamine in mice. Perindopril (0.05 and 0.1 mg/kg, PO) was given orally for one week before administration of scopolamine (3mg/kg, IP). Then, memory function was evaluated by Morris water maze and passive avoidance tests. CBF was measured by laser Doppler flowmetry. Biochemical and molecular parameters were estimated after the completion of behavioral studies. Scopolamine caused impairment in memory which was associated with reduced CBF, acetylcholine (ACh) level and elevated acetylcholinesterase (AChE) activity and malondialdehyde (MDA) level. Perindopril ameliorated scopolamine induced amnesia in both the behavioral paradigms. Further, perindopril prevented elevation of AChE and MDA level in mice brain. There was a significant increase in CBF and ACh level in perindopril treated mice. However, scopolamine had no significant effect on ATP level and mRNA expression of angiotensin receptors and ACE in cortex and hippocampus. But, perindopril significantly decreased ACE activity in brain without affecting its mRNA expression. The study clearly showed the interaction between ACE and cholinergic neurotransmission and beneficial effect of perindopril can be attributed to improvement in central cholinergic neurotransmission and CBF., (Copyright © 2012. Published by Elsevier B.V.)

Introduction: Pioglitazone, a peroxisome proliferator activated receptor γ (PPARγ) agonist, is widely used in clinical medicine as a treatment for type 2 diabetes and is recently proved to have beneficial effects on improving cognition in early stages of Alzheimer's disease (AD). Moreover, it has been shown that pioglitazone reduces N-methyl-D-aspartate (NMDA, a glutamate agonist) mediated calcium currents and transients. Since enhanced calcium transients are present in AD models, we tested the hypothesis whether pioglitazone manifests its acquisition memory enhancement role through glutamatergic pathway., Material and Methods: Memory performance was evaluated in a two-trial recognition Y-maze test and passive avoidance in mice. Pioglitazone (20 or 40 mg/kg, p.o.) was administered 2h before each trial, NMDA (75 mg/kg i.p.), 15 min before pioglitazone, and scopolamine, an M1 (muscarinic) receptor antagonist (0.3 or 1.0 mg/kg i.p.) and MK-801 (dizocilpine) (0.01, 0.03 or 0.1 mg/kg, i.p.), the highly selective, non-competitive NMDA antagonist--30 min beforehand., Results: (1) We induced the memory impairment by scopolamine or MK-801 before trials. (2) Pioglitazone did not improve the memory impairment induced by MK-801. (3) Pioglitazone significantly improved the memory impairment induced by scopolamine. (4) Subeffective dose of MK-801 nullified the beneficial effects of pioglitazone in scopolamine induced memory impaired mice. (5) NMDA promoted the effects of subeffective dose of pioglitazone on memory impaired by scopolamine., Discussion: In conclusion, the present study suggests that glutamatergic pathway is involved in the pioglitazone induced memory performance., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Cocaine addiction is a public health issue in many countries, stressing the need for more effective treatments. As all drugs of abuse, cocaine acts on the brain reward system, increasing dopamine (DA) levels. Other neurotransmitters such as acetylcholine (ACh) are involved in the mechanisms underlying the development and the maintenance of cocaine addiction. ACh plays an important role in learning and memory processes and also regulates DA in some specific regions of the central nervous system. The present study investigated the effects of biperiden, a muscarinic cholinergic (mACh) antagonist in two animal models: conditioned place preference (CPP) and behavioral sensitization. Male C57BL/6J mice were used in both studies. The CPP protocol was unbiased and carried out in three phases: habituation, conditioning and testing. For conditioning, cocaine was injected at a dose of 10mg/kg in eight 15 min-sessions. The treatment with biperiden (doses of 0.1, 1 and 10 mg/kg) was made 30 min prior to the testing session. For behavioral sensitization development, cocaine was administered at the dose of 10 mg/kg for 10 days. After sensitization, two challenges were performed: saline and cocaine (5 mg/kg). Biperiden (10 mg/kg) was administered 30 min before the cocaine challenge. At the dose of 10 mg/kg, biperiden blocked the cocaine-CPP expression, suggesting an effect on conditioned memory retrieval. However, the same dose potentiated the expression of behavioral sensitization, suggesting an increase in DA release, probably in the NAc. Biperiden, as other mACh antagonists, may be a promising drug for the pharmacologic treatment of cocaine addiction., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Bile duct ligation (BDL) induces primary biliary cirrhosis characterized by cholestasis, impaired liver function and cognition including impairment of memory formation and anxiety-like behaviors. Endogenous opioid and acetylcholine levels are elevated in animal model of cholestasis. In addition, there is no data about the effects of interaction opioidergic and cholinergic systems of dorsal hippocampus (CA1) on amnesia-induced by cholestasis. Male mice weighing 25-35 g were used in this study. Cholestasis was induced by the ligation of the common bile duct. One-trial step-down and hole-board paradigms were used for the assessment of memory retrieval and anxiety-like behaviors respectively. All drugs injected intra-CA1. The data showed that cholestasis (24 days after BDL) decreased memory retrieval. Sole intra-CA1 injection of higher dose of mecamylamine (0.125, 0.25, 0.5 and 1 μg/mice) and scopolamine (0.125, 0.25, 0.5 1 and 2 μg/mice) but not all doses of naloxone (0.0125, 0.025 and 0.05 μg/mice) decreased memory retrieval in the sham operated BDL. The ineffective doses of naloxone (0.025 and 0.05 μg/mice), mecamylamine (0.5 μg/mice) and scopolamine (0.5 μg/mice) restored cholestasis-induced amnesia 24 days after BDL. Further, all cross co-administration ineffective doses of naloxone (0.0125 μg/mice), mecamylamine (0.125 μg/mice) and scopolamine (0.125 μg/mice) reversed cholestasis-induced amnesia. All doses of the drugs have no effect on exploratory behaviors. The data strongly revealed that synergistic effect between opioidergic and cholinergic systems of CA1 on the modulation of cholestasis-induced amnesia., (Copyright © 2011 Elsevier B.V. All rights reserved.)

DYT1 early-onset generalized torsion dystonia is an inherited movement disorder associated with mutations in DYT1 that codes for torsinA protein. The most common mutation seen in this gene is a trinucleotide deletion of GAG. We previously reported a motor control deficit on a beam-walking task in our Dyt1 ΔGAG knock-in heterozygous mice. In this report we show the reversal of this motor deficit with the anticholinergic trihexyphenidyl (THP), a drug commonly used to treat movement problems in dystonia patients. THP also restored the reduced corticostriatal long-term depression (LTD) observed in these mice. Corticostriatal LTD has long been known to be dependent on D2 receptor activation. In this mouse model, striatal D2 receptors were expressed at lower quantities in comparison to wild-type mice. Furthermore, the mice were also partially resistant to FPL64176, an agonist of L-type calcium channels that have been previously reported to cause severe dystonic-like symptoms in wild-type mice. Our findings collectively suggest that altered communication between cholinergic interneurons and medium spiny neurons is responsible for the LTD deficit and that this synaptic plasticity modification may be involved in the striatal motor control abnormalities in our mouse model of DYT1 dystonia., (Copyright © 2011 Elsevier B.V. All rights reserved.)

This study presents an in vivo investigation of the arylpropylsulfonamide α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptor positive modulator (R,R)-N,N-(2,20-[biphenyl-4-40-diyl]bis[propane-2,1-diyl])dimethanesulfonamide (PIMSD). The pharmacokinetics of the drug were examined in male C57BL/6J mice and the drug concentration in blood plasma determined after subcutaneous injection of 1mg/kg b.w. This analysis revealed a rapid increase of the plasma concentration, peaking within 30min after administration with a T(1/2) of approximately 30min and a peak plasma concentration of about 2μM. Analysis of brain tissue homogenates also indicated blood-brain barrier permeability of the compound. Cognitive enhancing effects of the drug were then studied on place learning in male C57BL/6J mice in a water maze. In order to elucidate the potential positive effects of PIMSD on spatial learning the muscarinergic antagonist scopolamine was utilized, which is known to impair spatial learning ability. The mice were divided into four groups and subjected to two sequential subcutaneous injections administered 25min prior to behavioural testing: (1) vehicle/vehicle; (2) PIMSD/vehicle; (3) scopolamine/vehicle; (4) PIMSD/scopolamine. PIMSD at a dose of 3mg/kg b.w. was able to partially reverse the impairment given by 0.5mg/kg b.w. scopolamine. These results suggest that arylpropylsulfonamides such as PIMSD may have a therapeutic use in the enhancement of cognitive function and support the hypothesis that AMPA receptor potentiation is one mechanism that can be targeted for diseases of cognitive impairment., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Biperiden is one of the most commonly abused anticholinergic drugs. This study assessed its motivational effects in the acquisition of conditioned place preference in rats. Biperiden neither produced place conditioning itself nor enhanced the rewarding effect of morphine. Furthermore, biperiden in combination with haloperidol also did not affect place preference. These findings suggest that biperiden seems devoid of abuse potential properties at least at the doses used., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Retrieval of memory is fundamental for our life as individuals. The participation of cholinergic system in memory consolidation process has been extensively studied, but there are few data concerning the function of this system in memory retrieval process. In the current study, we inject non-selective muscarinic antagonist scopolamine peripherally 20 min before training or testing to see whether cholinergic modulation has effects on the acquisition or retrieval of spatial memory by water maze task and fear memory by inhibitory avoidance task. We find that the cholinergic system is essential for the acquisition of both spatial memory and fear memory. As for the memory retrieval, the cholinergic system has a positive role in the retrieval of spatial memory, because mice injected with scopolamine 20 min before the testing in the water maze show impaired spatial memory retrieval. Whereas injection of scopolamine 20 min before the testing in the inhibitory avoidance task does not cause memory retrieval deficits. That indicates the cholinergic system is not essential for the retrieval of fear memory., (Copyright © 2010 Elsevier B.V. All rights reserved.)

To explore spatial cognition in rodents, research uses maze tasks, which differ in complexity, number of goals and pathways, behavioural flexibility, memory duration, but also in the experimenter's control over the strategy developed to reach a goal (e.g., allocentric vs. egocentric). This study aimed at validating a novel spatial memory test: the double-H maze test. The transparent device made of an alley with two opposite arms at each extremity and two in its centre is flooded. An escape platform is submerged in one arm. For experiments 1-3, rats were released in unpredictable sequences from one of both central arms to favour an allocentric approach of the task. Experiment 1 (3 trials/day over 6 days) demonstrated classical learning curves and evidence for recent and nondegraded remote memory performance. Experiment 2 (2 days, 3 trials/day) showed a dose-dependent alteration of task acquisition/consolidation by muscarinic or NMDA receptor blockade; these drug effects vanished with sustained training (experiment 3; 4 days, 3 trials/day). Experiment 4 oriented rats towards a procedural (egocentric) approach of the task. Memory was tested in a misleading probe trial. Most rats immediately switched from response learning-based to place learning-based behaviour, but only when their initial view on environmental cues markedly differed between training and probe trials. Because this simple task enables the formation of a relatively stable memory trace, it could be particularly adapted to study consolidation processes at a system level or/and the interplay between procedural and declarative-like memory systems., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Numerous studies have demonstrated cognitive improvements resulting from the application of nicotine, especially in those tasks aimed at measuring attention. While the neuro-pharmacological relationship between nicotine and acetylcholine-driven attentional processes has been examined, studies tend to focus on the duration of time in which a subject can attend to a specific stimulus or series of stimuli rather than on the subjects' adaptive attentional capabilities. The present study addresses the possibility that the cholinergic agonist nicotine could improve performance on a task testing the ability to shift attention between sensory modalities under both normal and pharmacologically impaired conditions. In a pilot set of experiments, we tested the effects of nicotine in a cross-modal experimental task designed to tax both the auditory and visual systems of male Sprague-Dawley rats. Nicotine (0.2 mg/kg) significantly improved performance on both auditory and visual trials, under repetitive trial conditions, and significantly decreased overall response latency. For the primary study, we tested the effects of decreasing cholinergic neurotransmission by systemic administration of the muscarinic antagonist atropine. Atropine (12.5 mg/kg) significantly impaired performance in auditory shift trials and perseverative trials, while significantly increasing the overall response latency. We then tested the effect of nicotine within the impaired model. Systemic administration of nicotine significantly improved performance in auditory and visual shift trials, while showing moderate improvements in response latency and perseverative trial conditions. These results indicate the potential therapeutic use of nicotine as a cognitive enhancer, as well as provide evidence for cholinergic system compensations., (Published by Elsevier B.V.)

The cGMP/PK-G pathway plays a crucial role in neuroprotection and neurotrophin support, and is possibly involved in antidepressant action. Recently we reported on a novel antidepressant-like response following simultaneous administration of sildenafil (phosphodiesterase 5 (PDE5) inhibitor, thereby increasing cGMP levels), and atropine (muscarinic acetylcholine receptor antagonist) in the rat forced swim test (FST). However, it is unclear whether the antidepressant-like activity of sildenafil+atropine is mediated via the activation of PK-G, an important down-stream effector for cGMP, and whether this may target known pathways in antidepressant action. We investigated whether the antidepressant-like response of sildenafil+/-atropine could be reversed by Rp-8-Br-PET-cGMP, a PK-G inhibitor, and also whether a combination of 8-Br-cGMP (PK-G activator)+/-atropine would likewise be active in the FST, and whether this combination could be attenuated by a PK-G inhibitor. 8-Br-cGMP alone, but not sildenafil alone, reduced immobility and selectively increased swimming in the FST. The antidepressant-like action of sildenafil was only evident following co-administration of atropine, and selectively increased climbing behaviour. Importantly, PK-G inhibition prevented the antidepressant-like effects of both 8-Br-cGMP and the sildenafil/atropine combination. These results confirm cholinergic-cGMP-PK-G interactions in the antidepressant-like effects of sildenafil, putatively acting via noradrenergic mechanisms, whereas direct PK-G activation induces antidepressant-like effects that are associated with enhancement of serotonergic neurotransmission.

Decrease in the expression or activity of acetylcholinesterase (AChE) enzymatic activity results in increased cholinergic tonus in the brain and periphery, with concomitant regulations of nicotinic and muscarinic receptors expression. We generated AChE knockout mice and characterized the behavioral phenotype of heterozygous animals, focusing on learning and memory functions. Male and female, AChE+/- and AChE+/+ littermate controls (129 sv strain) were tested at 5-9 weeks of age. AChE activity was significantly decreased in the hippocampus and cortex of AChE+/- mice, but butyrylcholinesterase activity was preserved. AChE+/- mice failed to show any difference in terms of locomotion, exploration and anxiety parameters in the open-field test. Animals were then tested for place learning in the water-maze. They were trained using a 'sustained acquisition' protocol (3 swim trials per day) or a 'mild acquisition' protocol (2 swim trials per day) to locate an invisible platform in fixed position (reference memory procedure). Then, during 3 days, they were trained to locate the platform in a variable position (working memory procedure). Learning profiles and probe test performances were similar for AChE+/- and AChE+/+ mice. Mice were then treated with the muscarinic receptor antagonist scopolamine (0.5, 5 mg/kg) 20 min before each training session. Scopolamine impaired learning at both doses in AChE+/+ mice, but only at the highest dose in AChE+/- mice. Moreover, the intracerebroventricular injection of amyloid-beta25-35 peptide, 9 nmol, 7 days before water-maze acquisition, failed to induce learning deficits in AChE+/- mice, but impaired learning in AChE+/+ controls. The peptide failed to be toxic in forebrain structures of AChE+/- mice, since an increase in lipid peroxidation levels was measured in the hippocampus of AChE+/+ but not AChE+/- mice. We conclude that the increase in cholinergic tonus observed in AChE+/- mice did not result in increased memory functions but allowed a significant prevention of the deleterious effects of muscarinic blockade or amyloid toxicity.

Symptoms of schizophrenia, commonly divided into positive symptoms, negative symptoms, and cognitive impairments, exhibit different sensitivity to pharmacological treatments. As such, they are typically modeled in animals by behavioral effects of drugs that evoke these symptoms in humans, such as amphetamine or phencyclidine (PCP). Despite the fact that muscarinic antagonists also evoke a schizophrenia-like syndrome ("antimuscarinic syndrome") and findings of cholinergic-related alterations in brains of schizophrenia patients, modeling schizophrenia using muscarinic manipulations has been infrequently considered, and the effects of muscarinic blockade on behavioral tasks relevant to schizophrenia have not been adequately characterized. The present review surveys recent attempts to model schizophrenia-related symptoms using manipulations causing cholinergic dysfunction, particularly muscarinic blockade, in well validated behavioral models of schizophrenia, such as prepulse inhibition and latent inhibition.

Object location task (OLT) has been used as a model of hippocampal-dependent memory. Despite this application, there is neither a consistent pharmacological validation of NMDA receptor modulation nor an evaluation of hippocampal participation in mice. In the OLT, mice were placed in the open field with two identical objects for 3 min and, after a delay of 30, 90, 180 or 360 min, one object was moved to a new location and the time spent exploring the objects in new, (novel) and old (familiar) locations was recorded. Our results showed that the mice were able to discriminate object location when tested either 90 or 180 min after training. Intraperitoneal administration of MK801 (NMDA receptors antagonist) or scopolamine (mACh antagonist) induced amnesic effects. On the other hand, D-cycloserine (NMDA agonist) or tacrine (cholinesterase inhibitor) were able to improve memory in the mice tested. In addition, lidocaine infusion in the hippocampal CA1 region 10 min before training blocked object location memory. In short, this work indicates that OLT is susceptible to modulation of NMDA receptors, cholinergic neurotransmission and it is the first to characterize the participation of the hippocampal CA1 region, in this task.

Deficits in social behaviour is a characteristic of numerous mental disorders including autism, schizophrenia, depression and Alzheimer's disease. For the assessment of pharmacological and genetic experimental disease models, conventional social interaction tasks bear the uncertainty that any drug-induced abnormality of the investigator may feed back to the drug-free companion modifying its reactions. A considerable technical improvement was recently reported by Moy et al. [Moy SS, Nadler JJ, Perez A, Barbaro RP, Johns JM, Magnuson T, et al. Sociability and preference for social novelty in five inbred strains: an approach to assess autistic-like behaviours in mice. Genes Brain Behav 2004;3:287-302] in which the drug free partner is confined to a small cage and social contacts of the investigator are recorded uncontaminated of any social reactions of the stranger. Using this novel behavioural paradigm, we here show in C57Bl/6 female mice that sociability (social interaction with a stranger mouse) is not impaired after administration of the anxiolytic diazepam (0.1-1 mg/kg) or the muscarinic antagonist scopolamine hydrobromide (0.1-1 mg/kg). However, social memory tested after a short time interval was impaired by both drugs in a dose-dependent manner (diazepam: > or = 0.5mg/kg; scopolamine: > or = 0.3mg/kg). The scopolamine-induced short-term memory deficit was reversed to normal by the choline esterase inhibitor donepezil (1 mg/kg). Given this dependence of social recognition on the cholinergic system, combined with the clinical observation of reduced social contacts in dementia patients, sociability may offer a novel endpoint biomarker with translational value in experimental models of cognitive dysfunction.