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Repeated forced swim (FS) conditioning enhances nociceptive responses to temporomandibular joint (TMJ) stimulation in female rats. The basis for FS-induced TMJ hyperalgesia remains unclear. To test the hypothesis that serotonin 3 receptor (5HT3R) mechanisms contribute to enhanced TMJ nociception after FS, ovariectomized female rats were treated with estradiol and subjected to FS for three days. On day 4, rats were anesthetized with isoflurane and TMJ-responsive neurons were recorded from superficial and deep laminae at the trigeminal subnucleus caudalis/upper cervical (Vc/C1-2) region and electromyographic (EMG) activity was recorded from the masseter muscle. Only Vc/C1-2 neurons activated by intra-TMJ injections of ATP were included for further analysis. Although neurons in both superficial and deep laminae were activated by ATP, only neurons in deep laminae displayed enhanced responses after FS. Local application of the 5HT3R antagonist, ondansetron (OND), at the Vc/C1-2 region reduced the ATP-evoked responses of neurons in superficial and deep laminae and reduced the EMG response in both sham and FS rats. OND also decreased the spontaneous firing rate of neurons in deep laminae and reduced the high-threshold convergent cutaneous receptive field area of neurons in superficial and deep laminae in both sham and FS rats. These results revealed that central application of a 5HT3R antagonist, had widespread effects on the properties of TMJ-responsive neurons at the Vc/C1-2 region and on jaw muscle reflexes under sham and FS conditions. It is concluded that 5HT3R does not play a unique role in mediating stress-induced hyperalgesia related to TMJ nociception., (Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.)

Dry eye (DE) disease is commonly associated with ocular surface inflammation, an unstable tear film and symptoms of irritation. However, little is known about the role of central neural mechanisms in DE. This study used a model for persistent aqueous tear deficiency, exorbital gland removal, to assess the effects of mustard oil (MO), a transient receptor potential ankyrin (TRPA1) agonist, on eyeblink and eyewipe behavior and Fos-like immunoreactivity (Fos-LI) in the trigeminal brainstem of male rats. Spontaneous tear secretion was reduced by about 50% and spontaneous eyeblinks were increased more than 100% in DE rats compared to sham rats. MO (0.02-0.2%) caused dose-related increases in eyeblink and forelimb eyewipe behavior in DE and sham rats. Exorbital gland removal alone was sufficient to increase Fos-LI at the ventrolateral pole of trigeminal interpolaris/caudalis (Vi/Vc) transition region, but not at more caudal regions of the trigeminal brainstem. Under barbiturate anesthesia ocular surface application of MO (2-20%) produced Fos-LI in the Vi/Vc transition, in the mid-portions of Vc and in the trigeminal caudalis/upper cervical spinal cord (Vc/C1) region that was significantly greater in DE rats than in sham controls. MO caused an increase in Fos-LI ipsilaterally in superficial laminae at the mid-Vc and Vc/C1 regions in a dose-dependent manner. Smaller, but significant, increases in Fos-LI also were seen in the contralateral Vc/C1 region in DE rats. TRPA1 protein levels in trigeminal ganglia from DE rats ipsilateral and contralateral to gland removal were similar. Persistent tear reduction enhanced the behavioral and trigeminal brainstem neural responses to ocular surface stimulation by MO. These results suggested that TRPA1 mechanisms play a significant role in the sensitization of ocular-responsive trigeminal brainstem neurons in this model for tear deficient DE., (Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.)

Cornea-evoked eyeblinks maintain tear film integrity on the ocular surface in response to dryness and protect the eye from real or potential damage. Eyelid movement following electrical stimulation has been well studied in humans and animals; however, the central neural pathways that mediate protective eyeblinks following natural nociceptive signals are less certain. The aim of this study was to assess the role of the trigeminal subnucleus interpolaris/caudalis (Vi/Vc) transition and subnucleus caudalis/upper cervical cord (Vc/C1) junction regions on orbicularis oculi electromyographic (OOemg) activity evoked by ocular surface application of hypertonic saline or exposure to bright light in urethane anesthetized male rats. The Vi/Vc and Vc/C1 regions are the main sites of termination for trigeminal afferent nerves that supply the ocular surface, while hypertonic saline (saline=0.15-5M) and bright light (light=5k-20klux) selectively activate ocular surface and intraocular trigeminal nerves, respectively, and excite second-order neurons at the Vi/Vc and Vc/C1 regions. Integrated OOemg activity, ipsilateral to the applied stimulus, increased with greater stimulus intensities for both modalities. Lidocaine applied to the ocular surface inhibited OOemg responses to hypertonic saline, but did not alter the response to light. Lidocaine injected into the trigeminal ganglion blocked completely the OOemg responses to hypertonic saline and light indicating a trigeminal afferent origin. Synaptic blockade by cobalt chloride of the Vi/Vc or Vc/C1 region greatly reduced OOemg responses to hypertonic saline and bright light. These data indicate that OOemg activity evoked by natural stimuli known to cause irritation or discomfort in humans depends on a relay in both the Vi/Vc transition and Vc/C1 junction regions., (Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.)

Sensory input from the temporomandibular joint (TMJ) to neurons in superficial laminae at the spinomedullary (Vc/C1-2) region is strongly influenced by estrogen status. This study determined if GABAergic mechanisms play a role in estrogen modulation of TMJ nociceptive processing in ovariectomized female rats treated with high- (HE) or low-dose (LE) estradiol (E2) for 2days. Superficial laminae neurons were activated by ATP (1mM) injections into the joint space. The selective GABAA receptor antagonist, bicuculline methiodide (BMI, 5 or 50μM, 30μl), applied at the site of recording greatly enhanced the magnitude and duration of ATP-evoked responses in LE rats, but not in units from HE rats. The convergent cutaneous receptive field (RF) area of TMJ neurons was enlarged after BMI in LE but not HE rats, while resting discharge rates were increased after BMI independent of estrogen status. By contrast, the selective GABAA receptor agonist, muscimol (50μM, 30μl), significantly reduced the magnitude and duration of ATP-evoked activity, resting discharge rate, and cutaneous RF area of TMJ neurons in LE and HE rats, whereas lower doses (5μM) affected only units from LE rats. Protein levels of GABAA receptor β3 isoform at the Vc/C1-2 region were similar for HE and LE rats. These results suggest that GABAergic mechanisms contribute significantly to background discharge rates and TMJ-evoked input to superficial laminae neurons at the Vc/C1-2 region. Estrogen status may gate the magnitude of GABAergic influence on TMJ neurons at the earliest stages of nociceptive processing at the spinomedullary region., (Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.)

Enhanced light sensitivity is a common feature of many neuro-ophthalmic conditions and some chronic headaches. Previously we reported that the bright light-evoked increases in trigeminal brainstem neural activity and lacrimation depended on a neurovascular link within the eye (Okamoto et al., 2012). However, the supraspinal pathways necessary for these light-evoked responses are not well defined. To assess the contribution of the posterior hypothalamic area (PH), a brain region closely associated with control of autonomic outflow, we injected bicuculline methiodide (BMI), a GABAa receptor antagonist, into the PH and determined its effect on the encoding properties of ocular neurons at the ventrolateral trigeminal interpolaris/caudalis transition (Vi/Vc) and caudalis/upper cervical cord junction (Vc/C1) regions and on reflex lacrimation in male rats under isoflurane anesthesia. BMI markedly reduced light-evoked (>80%) responses of Vi/Vc and Vc/C1 neurons at 10 min with partial recovery by 50 min after injection. BMI also reduced (>35%) the convergent cutaneous receptive field area of Vi/Vc and Vc/C1 ocular neurons indicating that both intra-ocular and periorbital cutaneous inputs were affected by changes in PH outflow. Light-evoked lacrimation was reduced by >35% at 10 min after BMI, while resting mean arterial pressure increased promptly and remained elevated (>20 mmHg) throughout the 50-min post-injection period. These results suggested that PH stimulation, acting in part through increased sympathetic activity, significantly inhibited light- and facial skin-evoked activity of ocular neurons at the Vi/Vc and Vc/C1 region. These data provide further support for the hypothesis that autonomic outflow plays a critical role in mediating light-evoked trigeminal brainstem neural activity and reflex lacrimation., (Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.)

Work relating the mechanical states of articular cartilage chondrocytes to their biosynthetic responses is based on measurements in isolated cells or cells in explant samples removed from their natural in situ environment. Neither the mechanics nor the associated biological responses of chondrocytes have ever been studied in cartilage within a joint of a live animal, and no such measurements have ever been performed using physiologically relevant joint loading through muscular contractions. The purpose of this study was to design and apply a method to study the mechanics of chondrocytes in the exposed but fully intact knee of live animals, which was loaded near-physiologically through muscular contraction. In order to achieve this purpose, we developed an accurate and reliable method based on two-photon laser excitation microscopy. Near-physiological knee joint loading was achieved through controlled electrical activation of the knee extensor muscles that compress the articulating surfaces of the femur, tibia and patella. Accuracy of the system was assessed by inserting micro-beads of known dimensions into the articular cartilage of the mouse knee and comparing the measured volumes and diameters in the principal directions with known values of the beads. Accuracy was best in the plane perpendicular to the optical axis (average error = 1%) while it was slightly worse, but still excellent, along the optical axis (average error = 3%). Reliability of cell volume and shape measurements was 0.5% on average, and 2.9% in the worst-case-scenario. Pilot measurements of chondrocyte deformations upon sub-maximal muscular loading causing a mean articular contact pressure of 1.9 ± 0.2 MPa showed an "instantaneous" decrease in cell height (17 ± 4.5%) and loss of cell volume (22.3 ± 2.4%) that took minutes to recover upon deactivation of the knee extensor muscles., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Excessive discomfort after exposure to bright light often occurs after ocular injury and during headache. Although the trigeminal nerve is necessary for light-evoked discomfort, the mechanisms underlying this phenomenon, often referred to generally as photophobia, are not well defined. Quantitative Fos-like immunoreactivity (Fos-LI) was used to determine the pattern of neuronal activation in the caudal brainstem after bright light stimulation and, secondly, whether a neurovascular mechanism within the eye contributes to this response. Under barbiturate anesthesia, male rats were exposed to low (1 x 10(4) lx) or high intensity (2 x 10(4) lx) light delivered from a thermal neutral source for 30 min (30 s ON, 30 s OFF) and allowed to survive for 90 min. Intensity-dependent increases in Fos-LI were seen in laminae I-II at the trigeminal caudalis/cervical cord junction region (Vc/C1) and nucleus tractus solitarius (NTS). Fos-LI also increased at the trigeminal interpolaris/caudalis transition (Vi/Vc(vl)) and dorsal paratrigeminal (dPa5) regions independent of intensity. Intravitreal injection of norepinephrine greatly reduced light-evoked Fos-LI at the Vc/C1, dPa5 and NTS, but not at the Vi/Vc transition. Lidocaine applied to the ocular surface had no effect on Fos-LI produced in trigeminal brainstem regions. These results suggested that multiple regions of the caudal trigeminal brainstem complex integrate light-related sensory information. Fos-LI produced at the dPa5 and NTS, coupled with norepinephrine-induced inhibition, was consistent with the hypothesis that light-evoked activation of trigeminal brainstem neurons involves an intraocular neurovascular mechanism with little contribution from neurons that supply the ocular surface.

The influence of estradiol (E2) treatment on temporomandibular joint (TMJ) nociceptive processing in the caudal trigeminal sensory brain stem complex was assessed in ovariectomized female rats by quantitative Fos-immunoreactivity (Fos-LI). After 2 days of daily injections of high (HE2) or low (LE2) dose E2 rats were anesthetized and the small fiber excitant, mustard oil (MO, 0-20%), was injected into the TMJ and after 2 h brains were processed for Fos-LI. TMJ-evoked Fos-LI in laminae I-II at the trigeminal subnucleus caudalis/upper cervical cord (Vc/C1-2) junction and the dorsal paratrigeminal region (dPa5) was significantly greater in HE2 than LE2 rats, while Fos-LI produced at the ventral trigeminal interpolaris/caudalis transition region (Vi/Vc(vl)) was similar. E2 treatment also modified the influence of N-methyl-D-aspartate (NMDA) and AMPA receptor antagonists on TMJ-evoked Fos-LI. The NMDA antagonist, MK-801, dose-dependently reduced the Fos-LI response at the Vc/C1-2 junction in HE2 rats, while only high dose MK-801 was effective in LE2 rats. MK801 reduced equally the Fos-LI response at the Vi/Vc transition in both groups, while only minor effects were seen at the dPa5 region. The AMPA receptor antagonist, NBQX, reduced Fos-LI at the Vc/C(1-2) and Vi/Vc(vl) regions in HE2 rats, while only high dose NBQX was effective in LE2 rats. NBQX did not reduce Fos-LI at the dPa5 region in either group. These results suggest that estrogen status plays a significant role in TMJ nociceptive processing at the Vc/C1-2 junction mediated, in part, through ionotropic glutamate receptor-dependent mechanisms.

Sexual dimorphism in copulatory behavior results from organizational actions of sex steroids (permanent effects of sex steroids occurring during early development). Reproductive success depends not only on copulatory behavior, but also on mate choice, which is often sexually dimorphic as well. The clearest example is sexual partner preference: the preference of males for female sexual partners and females for males. Are organizational hormone actions responsible for sexual differentiation of sexual partner preference? The zebra finch (Taeniopygia guttata) is a potentially valuable species for addressing this question, because the birds form life-long socially monogamous pair bonds. In one experiment, both early estrogen treatment (injection with estradiol benzoate-EB-for the first 2 weeks posthatch) and unisex housing during juvenile development independently resulted in a preference for females over males in two-choice tests, and only females that experienced both EB treatment and unisex living were more likely than controls to pair with other females in colony tests. In a second experiment, females injected with an estrogen synthesis inhibitor for the first week posthatch preferred to spend time near females instead of males in two-choice tests, unlike control females. These experiments suggest that sexual partner preference may result from organizational hormone actions in this pair-bonding species. Possible neural mechanisms or sites that could underly hormonal organization of sexual partner preference in birds and mammals include the anterior hypothalamic/preoptic area, the corticomedial amygdala, and its avian homologue nucleus taeniae of the archistriatum, the septum, and peripheral sensory processes.

Effective process control can only be achieved through an understanding of the operating issues of the reaction. The development and use of effective and rugged analytical methods is necessary to monitor these parameters. The intent of this paper is to present some key analytical issues encountered in the synthesis of MK-0679, an LTD4 antagonist. In a key step of the compound's synthesis, a prochiral diester intermediate undergoes an enantioselective enzymatic hydrolysis (in the presence of Triton X-100) leading to the (S)-ester acid. Subsequent processing transforms the ester acid into the final product. The residual amount of the detergent in the final product, the rapid determination of the enzymatic activity and the optical purity of the final product emerged as key issues in the control of the reaction. As a solution, two techniques were utilized and are presented: flow injection analysis and HPLC.

Young rats subjected to bilateral lesions to one of 37 different brain sites were initially trained to traverse a narrow runway to reach a goal box containing water. The test involved blocking a portion of the runway with sawdust and determining whether the animals would burrow through the sawdust to gain access to the goal box. Lesions to 30 brain sites produced deficient performance on the sawdust-digging problem. This study, combined with earlier investigation of 11 additional brain sites, shows that 41 of 48 total brain sites are involved in this task. These findings suggest that underlying brain mechanisms include a nonspecific mechanism and several specific mechanisms, such as visuospatial-attentional, visual discrimination, vestibular-proprioceptive-kinesthetic discrimination, response flexibility, but much less place learning mechanisms.

Adult rats, previously subjected to bilateral frontal, parietal, occipital, frontocingulate or posterior cingulate ablations, were required to learn two discrete latch-box tasks (sliding a barrel-bolt to the right and elevating a hook). Those groups showing significant learning impairments had lesions to either the frontal cortex or frontocingulate cortex. The data suggest that the neocortical focus for latch-box deficits lies rostral to Level 8 on the Lashley brain diagram, while the frontocingulate focus for latch-box deficits lies within the prelimbic area.

Previous lesion studies suggest that the dorsal caudatoputamen (DCP), globus pallidus, ventrolateral thalamus (VLT), substantia nigra, ventral tegmental area, superior colliculus (SC), median raphe, and pontine reticular formation are components of the general learning system (GLS) of the rat brain. The current study attempted to determine whether bilateral lesions to two components of the GLS (DCP/VLT, DCP/SC or VLT/SC) would produce greater deterioration of learning ability than bilateral lesions to only one component (DCP, VLT or SC). In all combinations examined, a second lesion added to the first led to a significantly greater learning decrement on a series of spatial reversal problems than that associated with the first lesion alone. These results are compatible with the view that the foregoing structures are elements of the same functional system concerned either directly or indirectly with general learning ability.

Previous studies have shown that weanling rats with electrolytic lesions of the globus pallidus (GP), substantia nigra (SN), median raphe (MR) or pontine reticular formation (PRF) are deficient in learning a wide variety of laboratory tasks. The current study was designed to investigate whether this nonspecific learning deficiency is due to destruction of cells intrinsic to these subcortical regions or to fibers passing through these regions. Accordingly, neurotoxic lesions of the GP, SN, MR or PRF were made in weanling rats using ibotenic acid. Rats were subsequently required to learn a visual discrimination, a 3-cul maze and a nonvisual (incline plane) discrimination. While those groups with GP, SN or MR lesions showed significant deficits on all three problems, only the animals with GP lesions exhibited deficits comparable in magnitude to those associated with corresponding electrolytic lesions. Animals with lesions to the PRF were impaired only on the nonvisual discrimination. These results suggest that while destruction of neurons alone within the GP, SN or MR can produce a nonspecific learning impairment, the combined destruction of neurons and fibers of passage within the SN, MR or PRF produces a more profound learning deficit.

Marijuana use has been reported to spuriously elevate immunoreactive human chorionic gonadotropin (HCG) in the serum of patients with testicular germ cell tumors. To reinvestigate this finding, we measured serum HCG and delta 9-tetrahydrocannabinol (delta 9-THC) levels in 16 men known or suspected to be marijuana users. Eight of the serum samples had measurable levels of delta 9-THC, while eight did not. None contained immunoreactive HCG. The addition of delta 9-THC directly to pooled male serum had no effect on the HCG radioimmunoassay. These results indicate that marijuana does not artificially elevate serum HCG concentrations.

Young rats prepared with discrete bilateral lesions to the cerebral cortex, cingulate cortex, or dorsal hippocampus were required to learn a white-black discrimination, a 3-cul maze, and a nonvisual inclined plane discrimination. Only those rats with parietal lesions were impaired in acquiring all three habits. Those with occipitotemporal, frontocingulate, or posterior cingulate lesions were impaired on two habits, those with dorsal hippocampal lesions were impaired on one habit, and those with frontal (motor) cortical lesions failed to show any impairment. These results coupled with others suggest that the parietal cortex is unsurpassed in the wide range of learning and retention deficits which follows restricted neopallial lesions.

Recent studies suggest that lesions of the median raphe or pontine reticular formation in adult rats are associated with a nonspecific (generalized) learning impairment. The present study showed that lesions in the region of the median raphe or pontine reticular formation in 21 day old rats likewise produced a nonspecific learning impairment, as evidence by significant deficits on a visual discrimination, nonvisual incline plane discrimination, 3-cul maze, and three simple spatial problems. The finding that relatively large lateral pontomesencephalic lesions did not lead to deficient learning of any of these tasks indicates that lesion locus rather than lesion size is responsible for the production of the learning deficits observed in this study.

Rats previously trained to run from a start box, through a choice chamber, approach and displace a white card rather than a black card and enter a goal box in order to avoid footshock sustained damage to either the medial thalamus, occipital cortex or rostral caudoputamen. The retention test disclosed that the avoidance response was abolished by thalamic lesions, the visual discrimination response was abolished by occipital lesions and the card displacement response was abolished by caudatal lesions. These data demonstrate that this visual discrimination task can be fractionated into a least three discrete habits: One is related to the initiation of the response leading to the reward (incentive habit), the second is related to the location of the correct pathway to the reward (location habit) and the third is related to the response necessary to gain access to the reward (response habit).

This study examined the relationship between self-reports of marijuana and cigarette smoking, and the physiological measures of expired air carbon monoxide (CO) and saliva thiocyanate (SCN) in a sample of 1,130 seventh, ninth, tenth, eleventh, and twelfth graders. Subjects who reported marijuana smoking were likely to also report cigarette smoking. The correlation between self-reported marijuana smoking and SCN was negligible. The correlation between CO and self-reported marijuana smoking was statistically significant, but when the variance due to cigarette smoking was removed, this relationship also became negligible. The existence of a sizeable number of marijuana smokers in this sample of adolescents did not alter the correlation between CO and self-reports of cigarette smoking. However, in adult samples, where marijuana and cigarette smoking are less highly correlated, marijuana smoking could affect the relationship between CO and self-reported cigarette smoking. Carbon monoxide predicted self-reported cigarette smoking better than did saliva thiocyanate. There was an interaction between grade and the CO/cigarette smoking correlation. The correlations were generally higher in upper grades.

Recent lesion studies on young rats suggest that the components of the rodent's general learning system (GLS; a group of brain structures essential for normal acquisition of a wide range of laboratory tasks, include the regions of the caudatoputamen, globus pallidus, ventrolateral thalamus, substantia nigra, ventral tegmental area, superior colliculus, median raphe, and pontine reticular formation). The current study provides evidence that young GLS-lesioned rats, like mentally retarded humans, may be suffering from a disturbance in some superordinate ability (executive functioning) that controls the use of learning strategies in general and the transfer of learning in particular. Specifically, thirsty rats were initially trained to traverse a narrow runway to reach a goal box containing water. When a portion of the runway was blocked with sawdust, all of the sham-operated control rats succeeded in burrowing through the sawdust to gain access to the goal box, whereas most of our GLS-lesioned rats failed to do so even though they "knew how" to dig. Neocortically damaged rats showed a similar though significantly smaller deficit. Although other interpretations are possible, these data give tentative support to the view that this impairment in transfer reflects a defect in executive processing.