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Abstract The astrocyte-to-neuron lactate shuttle model entails that, upon glutamatergic neurotransmission, glycolytically derived pyruvate in astrocytes is mainly converted to lactate instead of being entirely catabolized in mitochondria. The mechanism of this metabolic rewiring and its occurrence in human brain are unclear. Here by using immunohistochemistry (4 brains) and imaging mass cytometry (8 brains) we show that astrocytes of the adult human neocortex and hippocampal formation express barely detectable amounts of mitochondrial proteins critical for performing oxidative phosphorylation (OXPHOS). These data are corroborated by queries of transcriptomes (107 brains) of neuronal versus non-neuronal cells fetched from the Allen Institute for Brain Science for genes coding for a much larger repertoire of entities contributing to OXPHOS, showing that human non-neuronal elements barely expressed mRNAs coding for such proteins. With less OXPHOS, human brain astrocytes are thus bound to produce more lactate to avoid interruption of glycolysis.

Abstract The estrus cycle in female rodents has been shown to affect a variety of physiological functions. However, little is known about its presumably thorough effect on auditory processing during the sleep–wake cycle and sleep deprivation. Vertex auditory evoked potentials (vAEPs) were evoked by single click tone stimulation and recorded during different stages of the estrus cycle and sleep deprivation performed in metestrus and proestrus in female rats. vAEPs showed a strong sleep-dependency, with the largest amplitudes present during slow wave sleep while the smallest ones during wakefulness. Higher amplitudes and longer latencies were seen in the light phase during all vigilance stages. The largest amplitudes were found during proestrus (light phase) while the shortest latencies were seen during estrus (dark phase) compared to the 2nd day diestrus baseline. High-amplitude responses without latency changes were also seen during metestrus with increased homeostatic sleep drive. More intense and faster processing of auditory information during proestrus and estrus suggesting a more effective perception of relevant environmental cues presumably in preparation for sexual receptivity. A 4-h sleep deprivation resulted in more pronounced sleep recovery in metestrus compared to proestrus without difference in delta power replacement suggesting a better tolerance of sleep deprivation in proestrus. Sleep deprivation decreased neuronal excitability and responsiveness in a similar manner both during metestrus and proestrus, suggesting that the negative consequences of sleep deprivation on auditory processing may have a limited correlation with the estrus cycle stage.

Summary: The medial preoptic area (MPOA) has long been implicated in maternal and male sexual behavior. Modern neuroscience methods have begun to reveal the cellular networks responsible, while also implicating the MPOA in other social behaviors, affiliative social touch, and aggression. The social interactions rely on input from conspecifics whose most important modalities in rodents are olfaction and somatosensation. These inputs bypass the cerebral cortex to reach the MPOA to influence the social function. Hormonal inputs also directly act on MPOA neurons. In turn, the MPOA controls social responses via various projections for reward and motor output. The MPOA thus emerges as one of the major brain centers for instinctive social behavior. While key elements of MPOA circuits have been identified, a synthesis of these new data is now provided for further studies to reveal the mechanisms by which the area controls social interactions.

(1) Background: The effects of short-term social isolation during adulthood have not yet been fully established in rats behaviourally, and not at all transcriptomically in the medial prefrontal cortex (mPFC). (2) Methods: We measured the behavioural effects of housing adult male rats in pairs or alone for 10 days. We also used RNA sequencing to measure the accompanying gene expression alterations in the mPFC of male rats. (3) Results: The isolated animals exhibited reduced sociability and social novelty preference, but increased social interaction. There was no change in their aggression, anxiety, or depression-like activity. Transcriptomic analysis revealed a differential expression of 46 genes between the groups. The KEGG pathway analysis showed that differentially expressed genes are involved in neuroactive ligand-receptor interactions, particularly in the dopaminergic and peptidergic systems, and addiction. Subsequent validation confirmed the decreased level of three altered genes: regulator of G protein signalling 9 (Rgs9), serotonin receptor 2c (Htr2c), and Prodynorphin (Pdyn), which are involved in dopaminergic, serotonergic, and peptidergic function, respectively. Antagonizing Htr2c confirmed its role in social novelty discrimination. (4) Conclusions: Social homeostatic regulations include monoaminergic and peptidergic systems of the mPFC.

Summary: Microglial cells were eliminated from the brain with sustained 3–4 weeks long inhibition of colony stimulating factor 1 receptor by Pexidartinib 3397 (PLX3397). The prepartum treated mice mothers did not feed their pups after parturition. The pups of mothers treated orally only in the postpartum period starting immediately after parturition showed reduced body weight by 15.5 ± 0.22 postnatal days as the treatment progressed without the mothers showing altered caring behaviors. The apparent weight gain of foster pups during a suckling bout was reduced in mother mice fed by PLX3397-containing diet and also in rat dams following sustained intracerebroventricular infusion of PLX3397 in a separate experiment suggesting that lactation was affected by the reduced number of microglia. Prolactin secretion and signaling were markedly reduced in PLX3397-treated mothers. The results suggest that microglial cells are required for prolactin secretion and lactation whereas maternal motivation may not be directly affected by microglia.

Dibenzylbutyrolactone-type lignans are phenolic compounds of medical importance. The purpose of the study was to determine the effects of two such lignans, arctigenin and trachelogenin on the motility of isolated rat ileum and obtain indications on their mechanism of action. They were isolated from Arctium lappa and Cirsium arvense, respectively, which have been used traditionally to treat gastrointestinal disorders. 1–1.5 cm long segments of distal ileum were obtained from adult male Wistar rats. The intestinal segments were suspended vertically in a well-aerated organ-bath according to Magnus mounting method. The intestinal motility was monitored for 30 min before treatment to obtain the baseline, followed by treatment with 1 µM, 10 µM, 20 µM and 40 µM concentrations of arctigenin and 0.5 µM, 1 µM, 10 µM and 20 µM of trachelogenin concentrations. The amplitude, tone, and period of spontaneous contractions were measured after 15 and 30 min of treatment. To investigate their mechanism of action, cholinergic, glutamatergic, adrenergic antagonists and compounds inhibiting nitric oxide synthase and L-type calcium channels were also tested. Arctigenin and trachelogenin decreased the frequency of contractions in a dose-dependent manner. At the concentration of 20 µM and 40 µM of trachelogenin and arctigenin, respectively, there was a marked alteration in spontaneous contraction pattern with an observable increase in the period time. This activity was comparable to 0.5 µM nifedipine (L-type calcium channel blocker) treatment. Our results demonstrate relaxant effect of arctigenin and trachelogenin on the ileum motility that may be mediated by L-type calcium ion channel blockade.

The hypothalamo-pituitary system developed in early vertebrates. Prolactin is an ancient vertebrate hormone released from the pituitary that exerts particularly diverse functions. The purpose of the review is to take a comparative approach in the description of prolactin, its secretion from pituitary lactotrophs, and hormonal functions. Since the reproductive and osmoregulatory roles of prolactin are best established in a variety of species, these functions are the primary subjects of discussion. Different types of prolactin and prolactin receptors developed during vertebrate evolution, which will be described in this review. The signal transduction of prolactin receptors is well conserved among vertebrates enabling us to describe the whole subphylum. Then, the review focuses on the regulation of prolactin release in mammals as we have the most knowledge on this class of vertebrates. Prolactin secretion in response to different reproductive stimuli, such as estrogen-induced release, mating, pregnancy and suckling is detailed. Reproduction in birds is different from that in mammals in several aspects. Prolactin is released during incubation in avian species whose regulation and functional significance are discussed. Little information is available on prolactin in reptiles and amphibians; therefore, they are mentioned only in specific cases to explain certain evolutionary aspects. In turn, the osmoregulatory function of prolactin is well established in fish. The different types of pituitary prolactin in fish play particularly important roles in the adaptation of eutherian species to fresh water environments. To achieve this function, prolactin is released from lactotrophs in hyposmolarity, as they are directly osmosensitive in fish. In turn, the released prolactin acts on branchial epithelia, especially ionocytes of the gill to retain salt and excrete water. This review will highlight the points where comparative data give new ideas or suggest new approaches for investigation in other taxa.

The expression of the recently identified neuropeptide, amylin, is restricted in rodents to the postpartum preoptic area and may play a role in the control of parental behaviours and food intake. These processes are substantially different between bird and rodent parents as birds do not lactate but often show biparental care of the offspring. To establish the presence and role of amylin in the bird brain, in the present study, we investigated the distribution of amylin in brains of adult male and female zebra finches in three different reproductive stages (i.e. paired without young, incubating eggs or provisioning nestlings) and in unpaired control birds living in same sex flocks. Amylin mRNA was identified in the hypothalamus of zebra finch by RT-PCR, which was also used to produce probes for in situ hybridisation. Subsequently, in situ hybridisation histochemistry was performed in brain sections, and the labelling signal was quantified and compared between the groups. Amylin showed a much wider brain distribution than that of rodents. A strong and, in some regions, sexually dimorphic label was found in the striatum and several brain regions of the social behavioural network in both males and females. Many regions responsible for the learning of birdsong also contained amylin-positive neurons, and some regions showed sex differences reflecting the fact that vocalisation is sexually dimorphic in the zebra finch: only males sing. Area X (Ar.X), a striatal song centre present only in males, was labelled in paired but not unpaired male. Ar.X, another song centre, the lateral part of the magnocellular nucleus of the anterior nidopallium (lMAN) also contained amylin and had higher amylin label in paired, as opposed to unpaired birds. The wider distribution of amylin in birds as compared to rodents suggests a more general role of amylin in social or other behaviours in avian species than in mammals. Alternatively, parental care in birds may be a more complex behavioural trait involving a wider set of brain regions. The sex differences in song centres, and the changes with reproductive status suggest a participation of amylin in social behaviours and related changes in the singing of males.

The state of therapeutic ketosis can be achieved by using the ketogenic diet (KD) or exogenous ketone supplementation. It was suggested previously that the adenosinergic system may be involved in the mediating effect of KD on suppressing seizure activity in different types of epilepsies, likely by means of adenosine A1 receptors (A1Rs). Thus, we tested in the present study whether exogenous ketone supplements (ketone ester: KE, 2.5 g/kg/day; ketone salt/KS + medium chain triglyceride/MCT: KSMCT, 2.5 g/kg/day) applied sub-chronically (for 7 days) by intragastric gavage can modulate absence epileptic activity in genetically absence epileptic Wistar Albino Glaxo/Rijswijk (WAG/Rij) rats. The number of spike-wave discharges (SWDs) significantly and similarly decreased after both KE and KSMCT treatment between 3rd and 7th days of gavage. Moreover, blood beta-hydroxybutyrate (βHB) levels were significantly increased alike after KE and KSMCT gavage, compared to control levels. The SWD number and βHB levels returned to the baseline levels on the first day without ketone supplementation. To determine whether A1Rs can modify ketone supplement-evoked changes in absence epileptic activity, we applied a non-pro-epileptic dose of a specific A1R antagonist DPCPX (1,3-dipropyl-8-cyclopentylxanthine) (intraperitoneal/i.p. 0.2 mg/kg) in combination with KSMCT (2.5 g/kg/day, gavage). As expected, DPCPX abolished the KSMCT-evoked decrease in SWD number. Thus, we concluded that application of exogenous ketone supplements may decrease absence epileptic activity in WAG/Rij rats. Moreover, our results suggest that among others the adenosinergic system, likely via A1Rs, may modulate the exogenous ketone supplements-evoked anti-seizure effects.

Transforming growth factor-βs (TGF-βs) regulate cellular proliferation, differentiation, and survival. TGF-βs bind to type I (TGF-βRI) and II receptors (TGF-βRII), which are transmembrane kinase receptors, and an accessory type III receptor (TGF-βRIII). TGF-β may utilize another type I receptor, activin-like kinase receptor (Alk1). TGF-β is neuroprotective in the middle cerebral artery occlusion (MCAO) model of stroke. Recently, we reported the expression pattern of TGF-β1-3 after MCAO. To establish how TGF-βs exert their actions following MCAO, the present study describes the induction of TGF-βRI, RII, RIII and Alk1 at 24 h, 72 h and 1 mo after transient 1 h MCAO as well as following 24 h permanent MCAO using in situ hybridization histochemistry. In intact brain, only TGF-βRI had significant expression: neurons in cortical layer IV contained TGF-βRI. At 24 h after the occlusion, no TGF-β receptors showed induction. At 72 h following MCAO, all four types of TGF-β receptors were induced in the infarct area, while TGF-βRI and RII also appeared in the penumbra. Most cells with elevated TGF-βRI mRNA levels were microglia. TGF-βRII co-localized with both microglial and endothelial markers while TGF-βRIII and Alk1 were present predominantly in endothels. All four TGF-β receptors were induced within the lesion 1 mo after the occlusion. In particular, TGF-βRIII was further induced as compared to 72 h after MCAO. At this time point, TGF-βRIII signal was predominantly not associated with blood vessels suggesting its microglial location. These data suggest that TGF-β receptors are induced after MCAO in a timely and spatially regulated fashion. TGF-β receptor expression is preceded by increased TGF-β expression. TGF-βRI and RII are likely to be co-expressed in microglial cells while Alk1, TGF-βRII, and RIII in endothels within the infarct where TGF-β1 may be their ligand. At later time points, TGF-βRIII may also appear in glial cells to potentially affect signal transduction via TGF-βRI and RII.

Transforming growth factor-βs (TGF-β1-3) are cytokines that regulate the proliferation, differentiation, and survival of various cell types. The present study describes the induction of TGF-β1-3 in the rat after focal ischemia at 3 h, 24 h, 72 h and 1 month after transient (1 h) or permanent (24 h) middle cerebral artery occlusion (MCAO) using in situ hybridization histochemistry and quantitative analysis. Double labeling with different markers was used to identify the localization of TGF-β mRNA relative to the penumbra and glial scar, and the types of cells expressing TGF-βs. TGF-β1 expression increased 3 h after MCAO in the penumbra and was further elevated 24 h after MCAO. TGF-β1 was present mostly in microglial cells but also in some astrocytes. By 72 h and 1 month after the occlusion, TGF-β1 mRNA-expressing cells also appeared in microglia within the ischemic core and in the glial scar. In contrast, TGF-β2 mRNA level was increased in neurons but not in astrocytes or microglial cells in layers II, III, and V of the ipsilateral cerebral cortex 24 h after MCAO. TGF-β3 was not induced in cells around the penumbra. Its expression increased in only a few cells in layer II of the cerebral cortex 24 h after MCAO. The levels of TGF-β2 and -β3 decreased at subsequent time points. Permanent MCAO further elevated the levels of all 3 subtypes of TGF-βs suggesting that reperfusion is not a major factor in their induction. TGF-β1 did not co-localize with either Fos or ATF-3, while the co-localization of TGF-β2 with Fos but not with ATF-3 suggests that cortical spreading depolarization, but not damage to neural processes, might be the mechanism of induction for TGF-β2. The results imply that endogenous TGF-βs are induced by different mechanisms following an ischemic attack in the brain suggesting that they are involved in distinct spatially and temporally regulated inflammatory and neuroprotective processes.

BackgroundGlutamate (Glu) and gamma-aminobutyric acid (GABA) transporters play important roles in regulating neuronal activity. Glu is removed from the extracellular space dominantly by glial transporters. In contrast, GABA is mainly taken up by neurons. However, the glial GABA transporter subtypes share their localization with the Glu transporters and their expression is confined to the same subpopulation of astrocytes, raising the possibility of cooperation between Glu and GABA transport processes.Methodology/principal findingsHere we used diverse biological models both in vitro and in vivo to explore the interplay between these processes. We found that removal of Glu by astrocytic transporters triggers an elevation in the extracellular level of GABA. This coupling between excitatory and inhibitory signaling was found to be independent of Glu receptor-mediated depolarization, external presence of Ca(2+) and glutamate decarboxylase activity. It was abolished in the presence of non-transportable blockers of glial Glu or GABA transporters, suggesting that the concerted action of these transporters underlies the process.Conclusions/significanceOur results suggest that activation of Glu transporters results in GABA release through reversal of glial GABA transporters. This transporter-mediated interplay represents a direct link between inhibitory and excitatory neurotransmission and may function as a negative feedback combating intense excitation in pathological conditions such as epilepsy or ischemia.

Summary: The role of preoptic GABAergic inhibitory neurons was addressed in parenting, anxiety and depression. Pup exposure and forced swimming resulted in similar c-Fos activation pattern in neurons expressing vesicular GABA transporter in the preoptic area with generally stronger labeling and different distributional pattern in females than in males. Chemogenetic stimulation of preoptic GABAergic cells resulted in elevated maternal motivation and caring behavior in females and mothers but aggression toward pups in males. Behavioral effects were the opposite following inhibition of preoptic GABAergic neurons suggesting their physiological relevance. In addition, increased anxiety-like and depression-like behaviors were found following chemogenetic stimulation of the same neurons in females, whereas previous pup exposure increased only anxiety-like behavior suggesting that not the pups, but overstimulation of the cells can lead to depression-like behavior. A sexually dimorphic projection pattern of preoptic GABAergic neurons was also identified, which could mediate sex-dependent parenting and associated emotional behaviors.

Identity tracking and instance segmentation are crucial in several areas of biological research. Behavior analysis of individuals in groups of similar animals is a task that emerges frequently in agriculture or pharmaceutical studies, among others. Automated annotation of many hours of surveillance videos can facilitate a large number of biological studies/experiments, which otherwise would not be feasible. Solutions based on machine learning generally perform well in tracking and instance segmentation; however, in the case of identical, unmarked instances (e.g., white rats or mice), even state-of-the-art approaches can frequently fail. We propose a pipeline of deep generative models for identity tracking and instance segmentation of highly similar instances, which, in contrast to most region-based approaches, exploits edge information and consequently helps to resolve ambiguity in heavily occluded cases. Our method is trained by synthetic data generation techniques, not requiring prior human annotation. We show that our approach greatly outperforms other state-of-the-art unsupervised methods in identity tracking and instance segmentation of unmarked rats in real-world laboratory video recordings.

This study reports the distribution of parathyroid hormone 2 receptor (PTH2R)-immunoreactive fibers in the hypothalamus using fluorescent amplification immunocytochemistry. The pattern of immunolabeling is strikingly similar to that of tuberoinfundibular peptide of 39 residues (TIP39), a peptide recently purified from bovine hypothalamus and proposed to be a ligand of the PTH2R based on pharmacological data. To investigate the anatomical basis of suggestions that TIP39 affects the secretion of several hypophysiotropic hormones we performed double-labeling studies and found that only somatostatin fibers contain PTH2R in the median eminence, which suggests that somatostatin release could be directly regulated via the PTH2R. However, several hypothalamic nuclei projecting to the median eminence contain a high density of both TIP39 and PTH2R fibers and terminals. We report here, that the PTH2R terminals also contain vesicular glutamate transporter−2, and suggest that TIP39 terminals are ideally positioned to modulate glutamatergic influences on hypophysiotropic neurons. [ABSTRACT FROM AUTHOR]

Abstract Slow wave activity (SWA) is a characteristic brain oscillation in sleep and quiet wakefulness. Although the cell types contributing to SWA genesis are not yet identified, the principal role of neurons in the emergence of this essential cognitive mechanism has not been questioned. To address the possibility of astrocytic involvement in SWA, we used a transgenic rat line expressing a calcium sensitive fluorescent protein in both astrocytes and interneurons and simultaneously imaged astrocytic and neuronal activity in vivo. Here we demonstrate, for the first time, that the astrocyte network display synchronized recurrent activity in vivo coupled to UP states measured by field recording and neuronal calcium imaging. Furthermore, we present evidence that extensive synchronization of the astrocytic network precedes the spatial build-up of neuronal synchronization. The earlier extensive recruitment of astrocytes in the synchronized activity is reinforced by the observation that neurons surrounded by active astrocytes are more likely to join SWA, suggesting causality. Further supporting this notion, we demonstrate that blockade of astrocytic gap junctional communication or inhibition of astrocytic Ca2+ transients reduces the ratio of both astrocytes and neurons involved in SWA. These in vivo findings conclusively suggest a causal role of the astrocytic syncytium in SWA generation.

Abstract Adaptation to motherhood includes maternal behaviour and lactation during the postpartum period. The major organizing centres of maternal behaviour and lactation are located in the hypothalamic medial preoptic area (MPOA) and the arcuate nucleus, respectively. Insulin-like growth factor I (IGF-I) is an effector of the growth hormone axis; however, its function in the brain is largely unexplored. We identified increased maternal IGF binding protein-3 (IGFBP-3) expression in preoptic rat microarray data and confirmed it by RT-PCR. In situ hybridization histochemistry showed markedly elevated IGFBP-3 expression in the MPOA and the arcuate nucleus in rat dams. Prolonged intracerebroventricular injection of IGF-I or antagonism of brain IGFBP-3 with an inhibitor (NBI-31772) using osmotic minipumps increased pup retrieval time, suggesting reduced maternal motivation. Suckling-induced prolactin release and pup weight gain were also suppressed by IGF-I, suggesting reduced lactation. In addition, IGF-I-induced tyrosine hydroxylase expression and its specific phosphorylation in tuberoinfundibular dopaminergic neurons suppress prolactin secretion. Thus, IGF-I may inhibit both behavioural and lactational alterations in mothers. Neurons in the MPOA and arcuate nuclei express IGFBP-3 during the postpartum period to neutralize IGF-I effects. IGFBP-3 can prevent the blockade of maternal behaviour and lactation exerted by IGF-I, suggesting a novel modulatory mechanism underlying the behavioural and hormonal effects during central maternal adaptations.

Gonadotropin releasing hormone (GnRH) neurons provide neuronal input to the preoptic area (POA) and the arcuate nucleus (Arc), two regions involved critically in the regulation of neuroendocrine functions and associated behaviors. These areas contain tyrosine hydroxylase immunoreactive (TH-IR) neurons, which play location-specific roles in the neuroendocrine control of both the luteinizing hormone and prolactin secretion, as well as, sexually motivated behaviors. Concerning changes in the activity of GnRH neurons and the secretion pattern of GnRH seen under the influence of rising serum estrogen levels and during lactation, we tested the hypothesis that the functional state of GnRH neurons is mediated via direct synaptic connections to TH-IR neurons in the POA and Arc. In addition, we examined putative changes of these inputs in lactating mice and in mothers separated from their pups. Confocal microscopic and pre-embedding immunohistochemical studies on ovariectomized mice treated with 17β-estradiol (OVX+E2) provided evidence for direct appositions and asymmetric synapses between GnRH-IR fiber varicosities and TH-IR neurons in the POA and the Arc. As TH co-localizes with kisspeptin (KP) in the POA, confocal microscopic analysis was continued on sections additionally labeled for KP. The TH-IR neurons showed a lower level of co-labeling for KP in lactating mice compared to OVX+E2 mice (16.1 ± 5% vs. 57.8 ± 4.3%). Removing the pups for 24 h did not alter significantly the KP production in TH-IR neurons (17.3 ± 4.6%). The mean number of GnRH-IR varicosities on preoptic and arcuate TH cells did not differ in the three animal models investigated. This study shows evidence that GnRH neurons provide direct synaptic inputs to POA and Arc dopaminergic neurons. The scale of anatomical connectivity with these target cells was unaltered during lactation indicating a maintained GnRH input, inspite of the altered hormonal condition.

The member of synthetic cathinone family, methylenedioxypyrovalerone (MDPV), is a frequently used psychoactive drug of abuse. The objective of our study was to determine the effect of MDPV (administered from the 8th to the 14th day of gestation) on the behavior of neonatal and adolescent mice, as well as its effect on maternal care. We measured maternal care (pup retrieval test, nest building), locomotor activity (open field test), and motor coordination (grip strength test) of dams, whereas on pups we examined locomotor activity at postnatal day 7 and day 21 (open field test) and motor coordination on day 21 (grip strength test). On fresh-frozen brain samples of the dams we examined the expression of two important peptides implicated in the regulation of maternal behavior and lactation: tuberoinfundibular peptide 39 (TIP39) mRNA in the thalamic posterior intralaminar complex, and amylin mRNA in the medial preoptic nucleus. We detected decreased birth rate and survival of offspring, and reduced maternal care in the drug-treated animals, whereas there was no difference between the motility of treated and control mothers. Locomotor activity of the pups was increased in the MDPV treated group both at 7 and 21 days of age, while motor coordination was unaffected by MDPV treatment. TIP39 and amylin were detected in their typical location but failed to show a significant difference of expression between the drug-treated and control groups. The results suggest that chronic systemic administration of the cathinone agent MDPV to pregnant mice can reduce birth rate and maternal care, and it also enhances motility (without impairment of motor coordination) of the offspring.