Transcriptional signatures of fentanyl use in the mouse ventral tegmental area.

Synthetic opioids such as fentanyl contribute to the vast majority of opioid-related overdose deaths, but fentanyl use remains broadly understudied. Like other substances with misuse potential, opioids cause lasting molecular adaptations to brain reward circuits, including neurons in the ventral tegmental area (VTA). The VTA contains numerous cell types that play diverse roles in opioid use and relapse; however, it is unknown how fentanyl experience alters the transcriptional landscape in specific subtypes. Here, we performed single nuclei RNA sequencing to study transcriptional programs in fentanyl-experienced mice. Male and female C57/BL6 mice self-administered intravenous fentanyl (1.5 µg/kg/infusion) or saline for 10 days. After 24 h abstinence, VTA nuclei were isolated and prepared for sequencing on the 10× platform. We identified different patterns of gene expression across cell types. In dopamine neurons, we found enrichment of genes involved in growth hormone signalling. In dopamine-glutamate-GABA combinatorial neurons, and some GABA neurons, we found enrichment of genes involved in Pi3k-Akt signalling. In glutamate neurons, we found enrichment of genes involved in cholinergic signalling. We identified transcriptional regulators for the differentially expressed genes in each neuron cluster, including downregulated transcriptional repressor Bcl6, and upregulated transcription factor Tcf4. We also compared the fentanyl-induced gene expression changes identified in mouse VTA with a published rat dataset in bulk VTA, and found overlap in genes related to GABAergic signalling and extracellular matrix interaction. Together, we provide a comprehensive picture of how fentanyl self-administration alters the transcriptional landscape of the mouse VTA that serves as the foundation for future mechanistic studies.

EZH2-dependent epigenetic reprogramming in the central nucleus of amygdala regulates adult anxiety in both sexes after adolescent alcohol exposure.

Alcohol use and anxiety disorders occur in both males and females, but despite sharing similar presentation and classical symptoms, the prevalence of alcohol use disorder (AUD) is lower in females. While anxiety is a symptom and comorbidity shared by both sexes, the common underlying mechanism that leads to AUD and the subsequent development of anxiety is still understudied. Using a rodent model of adolescent intermittent ethanol (AIE) exposure in both sexes, we investigated the epigenetic mechanism mediated by enhancer of zeste 2 (EZH2), a histone methyltransferase, in regulating both the expression of activity-regulated cytoskeleton-associated protein (Arc) and an anxiety-like phenotype in adulthood. Here, we report that EZH2 protein levels were significantly higher in PKC-δ positive GABAergic neurons in the central nucleus of amygdala (CeA) of adult male and female rats after AIE. Reducing protein and mRNA levels of EZH2 using siRNA infusion in the CeA prevented AIE-induced anxiety-like behavior, increased H3K27me3, decreased H3K27ac at the Arc synaptic activity response element (SARE) site, and restored deficits in Arc mRNA and protein expression in both male and female adult rats. Our data indicate that an EZH2-mediated epigenetic mechanism in the CeA plays an important role in regulating anxiety-like behavior and Arc expression after AIE in both male and female rats in adulthood. This study suggests that EZH2 may serve as a tractable drug target for the treatment of adult psychopathology after adolescent alcohol exposure.

Tesofensine, a novel antiobesity drug, silences GABAergic hypothalamic neurons.

Obesity is a major global health epidemic that has adverse effects on both the people affected as well as the cost to society. Several anti-obesity drugs that target GLP-1 receptors have recently come to the market. Here, we describe the effects of tesofensine, a novel anti-obesity drug that acts as a triple monoamine neurotransmitter reuptake inhibitor. Using various techniques, we investigated its effects on weight loss and underlying neuronal mechanisms in mice and rats. These include behavioral tasks, DeepLabCut videotaped analysis, electrophysiological ensemble recordings, optogenetic activation, and chemogenetic silencing of GABAergic neurons in the Lateral Hypothalamus (LH). We found that tesofensine induces a greater weight loss in obese rats than lean rats, while differentially modulating the neuronal ensembles and population activity in LH. In Vgat-ChR2 and Vgat-IRES-cre transgenic mice, we found for the first time that tesofensine inhibited a subset of LH GABAergic neurons, reducing their ability to promote feeding behavior, and chemogenetically silencing them enhanced tesofensine's food-suppressing effects. Unlike phentermine, a dopaminergic appetite suppressant, tesofensine causes few, if any, head-weaving stereotypy at therapeutic doses. Most importantly, we found that tesofensine prolonged the weight loss induced by 5-HTP, a serotonin precursor, and blocked the body weight rebound that often occurs after weight loss. Behavioral studies on rats with the tastant sucrose indicated that tesofensine's appetite suppressant effects are independent of taste aversion and do not directly affect the perception of sweetness or palatability of sucrose. In summary, our data provide new insights into the effects of tesofensine on weight loss and the underlying neuronal mechanisms, suggesting that tesofensine may be an effective treatment for obesity and that it may be a valuable adjunct to other appetite suppressants to prevent body weight rebound.

Alcohol potentiates multiple GABAergic inputs to dorsal striatum fast-spiking interneurons.

Parvalbumin-expressing dorsal striatal fast-spiking interneurons, comprising ∼1% of the total dorsal striatal neuronal population, are necessary for the expression of compulsive-like ethanol consumption mice. Fast-spiking interneurons are driven to fire by glutamatergic inputs derived primarily from the cortex. However, these neurons also receive substantial GABAergic input from two sources: the globus pallidus and the reticular nucleus of the thalamus. How ethanol modulates inhibitory input onto fast-spiking neurons is unclear and, more broadly, alcohol effects on GABAergic synaptic transmission onto GABAergic interneurons are understudied. Examining this, we found that acute bath application of ethanol (50 mM) potentiated GABAergic transmission from both the globus pallidus and the reticular nucleus of the thalamus onto fast-spiking interneurons in mouse of both sexes. This ethanol-induced potentiation required postsynaptic calcium and was not accompanied by a sustained change in presynaptic GABA release probability. Examining whether this ethanol effect persisted following chronic intermittent ethanol exposure, we found attenuated acute-ethanol potentiation of GABAergic transmission from both the globus pallidus and the reticular nucleus of the thalamus onto striatal fast-spiking interneurons. These data underscore the impact of ethanol on GABAergic signaling in the dorsal striatum and support the notion that ethanol may disinhibit the dorsolateral striatum.

Sex-specific differences in KCC2 localisation and inhibitory synaptic transmission in the rat hippocampus.

Sexual differentiation of the brain is influenced by testosterone and its metabolites during the perinatal period, when many aspects of brain development, including the maturation of GABAergic transmission, occur. Whether and how testosterone signaling during the perinatal period affects GABAergic transmission is unclear. Here, we analyzed GABAergic circuit functional markers in male, female, testosterone-treated female, and testosterone-insensitive male rats after the first postnatal week and in young adults. In the hippocampus, mRNA levels of proteins associated with GABA signaling were not significantly affected at postnatal day (P) 7 or P40. Conversely, membrane protein levels of KCC2, which are critical for determining inhibition strength, were significantly higher in females compared to males and testosterone-treated females at P7. Further, female and testosterone-insensitive male rats at P7 showed higher levels of the neurotrophin BDNF, which is a powerful regulator of neuronal function, including GABAergic transmission. Finally, spontaneous GABAergic currents in hippocampal CA1 pyramidal cells were more frequent in females and testosterone-insensitive males at P40. Overall, these results show that perinatal testosterone levels modulate GABAergic circuit function, suggesting a critical role of perinatal sex hormones in regulating network excitability in the adult hippocampus.

Alcohol dependence and withdrawal increase sensitivity of central amygdalar GABAergic synapses to the glucocorticoid receptor antagonist mifepristone in male rats.

Aberrant glucocorticoid signaling via glucocorticoid receptors (GR) plays a critical role in alcohol use disorder (AUD). Acute alcohol withdrawal and protracted abstinence in dependent rats are associated with increased GR signaling and changes in GR-mediated transcriptional activity in the rat central nucleus of the amygdala (CeA). The GR antagonist mifepristone decreases alcohol consumption in dependent rats during acute withdrawal and protracted abstinence. Regulation of CeA synaptic activity by GR is currently unknown. Here, we utilized mifepristone and the selective GR antagonist CORT118335 (both at 10 µM) as pharmacological tools to dissect the role of GR on GABA transmission in male, adult Sprague-Dawley rats using slice electrophysiology. We subjected rats to chronic intermittent alcohol vapor exposure for 5-7 weeks to induce alcohol dependence. A subset of dependent rats subsequently underwent protracted alcohol withdrawal for 2 weeks, and air-exposed rats served as controls. Mifepristone reduced the frequency of pharmacologically-isolated spontaneous inhibitory postsynaptic currents (sIPSC) in the CeA (medial subdivision) without affecting postsynaptic measures in all groups, suggesting decreased GABA release with the largest effect in dependent rats. CORT118335 did not significantly alter GABA transmission in naïve, but decreased sIPSC frequency in dependent rats. Similarly, mifepristone decreased amplitudes of evoked inhibitory postsynaptic potentials only in dependent rats and during protracted withdrawal. Collectively, our study provides insight into regulation of CeA GABAergic synapses by GR. Chronic ethanol enhances the efficiency of mifepristone and CORT118335, thus highlighting the potential of drugs targeting GR as a promising pharmacological avenue for the treatment of AUD.

Effects of lanthanum nitrate on behavioral disorder, neuronal damage and gene expression in different developmental stages of Caenorhabditis elegans.

Rare earth elements (REEs) are widely used in the industry, agriculture, biomedicine, aerospace, etc, and have been shown to pose toxic effects on animals, as such, studies focusing on their biomedical properties are gaining wide attention. However, environmental and population health risks of REEs are still not very clear. Also, the REEs damage to the nervous system and related molecular mechanisms needs further research. In this study, the L1 and L4 stages of the model organism Caenorhabditis elegans were used to evaluate the effects and possible neurotoxic mechanism of lanthanum(III) nitrate hexahydrate (La(NO3)3·6H2O). For the L1 and L4 stage worms, the 48-h median lethal concentrations (LC50s) of La(NO3)3·6H2O were 93.163 and 648.0 mg/L respectively. Our results show that La(NO3)3·6H2O induces growth inhibition and defects in behavior such as body length, body width, body bending frequency, head thrashing frequency and pharyngeal pumping frequency at the L1 and L4 stages in C. elegans. The L1 stage is more sensitive to the toxicity of lanthanum than the L4 stage worms. Using transgenic strains (BZ555, EG1285 and NL5901), we found that La(NO3)3·6H2O caused the loss or break of soma and dendrite neurons in L1 and L4 stages; and α-synuclein aggregation in L1 stage, indicating that Lanthanum can cause toxic damage to dopaminergic and GABAergic neurons. Mechanistically, La(NO3)3·6H2O exposure inhibited or activated the neurotransmitter transporters and receptors (glutamate, serotonin and dopamine) in C. elegans, which regulate behavior and movement functions. Furthermore, significant increase in the production of reactive oxygen species (ROS) was found in the L4 stage C. elegans exposed to La(NO3)3·6H2O. Altogether, our data show that exposure to lanthanum can cause neuronal toxic damage and behavioral defects in C. elegans, and provide basic information for understanding the neurotoxic effect mechanism and environmental health risks of rare earth elements.

Learning-induced plasticity in the barrel cortex is disrupted by inhibition of layer 4 somatostatin-containing interneurons.

Gaba-ergic neurons are a diverse cell class with extensive influence over cortical processing, but their role in experience-dependent plasticity is not completely understood. Here we addressed the role of cortical somatostatin- (SOM-INs) and vasoactive intestinal polypeptide- (VIP-INs) containing interneurons in a Pavlovian conditioning where stimulation of the vibrissae is used as a conditioned stimulus and tail shock as unconditioned one. This procedure induces a plastic change observed as an enlargement of the cortical functional representation of vibrissae activated during conditioning. Using layer-targeted, cell-selective DREADD transductions, we examined the involvement of SOM-INs and VIP-INs activity in learning-related plastic changes. Under optical recordings, we injected DREADD-expressing vectors into layer IV (L4) barrels or layer II/III (L2/3) areas corresponding to the activated vibrissae. The activity of the interneurons was modulated during all conditioning sessions, and functional 2-deoxyglucose (2DG) maps were obtained 24 h after the last session. In mice with L4 but not L2/3 SOM-INs suppressed during conditioning, the plastic change of whisker representation was absent. The behavioral effect of conditioning was disturbed. Both L4 SOM-INs excitation and L2/3 VIP-INs inhibition during conditioning did not affect the plasticity or the conditioned response. We found the activity of L4 SOM-INs is indispensable in the formation of learning-induced plastic change. We propose that L4 SOM-INs may provide disinhibition by blocking L4 parvalbumin interneurons, allowing a flow of information into upper cortical layers during learning.

Moderate ethanol drinking is sufficient to alter Ventral Tegmental Area dopamine neurons activity via functional and structural remodeling of GABAergic transmission.

Earlier studies have shown a major involvement of Ventral Tegmental Area (VTA) dopamine (DA) neurons in mediating the rewarding effects of ethanol (EtOH). Much less is known on the role of this system in mediating the transition from moderate to excessive drinking and abuse. Here we sought to explore the hypothesis that early stage drinking in rodents, resembling recreational EtOH use in humans, is sufficient to dysregulate VTA DA transmission thus increasing the propensity to use over time. To this purpose, midbrain slice recordings in mice previously exposed to an escalating (3, 6 and 12%) 18-day voluntary EtOH drinking paradigm was used. By recording from DA and γ-aminobutyric acid (GABA) VTA neurons in midbrain slices, we found that moderate EtOH drinking leads to a significant suppression of the spontaneous activity of VTA DA neurons, while increasing their response to acute EtOH application. We also found that chronic EtOH leads to the enhancement of GABA input frequency onto a subset of DA neurons. Structurally, chronic EtOH induced a significant increase in the number of GABA axonal boutons contacting DA neurons, suggesting deep rewiring of the GABA network. This scenario is consistent with a downmodulation of the reward DA system induced by moderate EtOH drinking, a neurochemical state defined as "hypodopaminergic" and previously associated with advanced stages of drug use in humans. In this context, increased sensitivity of DA neurons towards acute EtOH may represent the neurophysiological correlate of increased unitary rewarding value, possibly driving progression to addiction.

The anxiolytic effect of salicylic acid is mediated via the GABAergic system in the fear potentiated plus maze behavior in rats.

BACKGROUND: Salicylic acid (SA) is a natural phenolic compound in plants with many beneficial effects for humans. The anxiolytic effect of this compound has been reported in animal models, but its mechanism of action remains unclear. In this study, by using the fear potentiated plus maze test, we evaluated the effect of salicylic acid on the gene expression of the main form of GABA (gamma aminobutyric acid) synthesizing enzyme i.e., the enzyme glutamic acid decarboxylase 67 (GAD67) which is called GAD1, in the ventral subiculum of the hippocampus, one of the main brain structures, in anxiety circuits. Also, the hypnotic effect of Salicylic acid was evaluated. METHODS: Animals were divided into the solvent, (SA) and diazepam treated groups (n = 6). For evaluating the anxiolytic effect of Salicylic acid, animals were subjected to 2 h of isolation, before placing them in the elevated plus maze (EPM). Afterward, the ventral part of the hippocampus was removed for evaluating the change in GAD1 gene expression by the reverse transcription-quantitative polymerase chain reaction (RTqPCR) technique. The hypnotic effect of Salicylic acid was evaluated in the ketamine induced sleeping test. RESULTS: Salicylic acid at 10 and 30 (mg/kg) increased time spent and entries to the open arms in the (EPM) (p < 0.05). (RTqPCR) revealed that 30 mg/kg of Salicylic acid increased GAD1 gene expression (p < 0.001). Salicylic acid (30 and 300 mg/kg) also increased the duration of sleep, in ketamine induced sleeping test (p < 0.05). CONCLUSION: Our results showed that Salicylic acid has anxiolytic and hypnotic effects and it exerts its anxiolytic effect partly, via up the regulation of GAD1 in the ventral part of the hippocampus.

Susceptibility to express amphetamine locomotor sensitization correlates with dorsolateral striatum bursting activity and GABAergic synapses in the globus pallidus.

Repeated psychostimulant administration results in behavioral sensitization, a process that is relevant in the early phases of drug addiction. Critically, behavioral sensitization is not observed in all subjects. Evidence shows that differential neuronal activity in the dorsolateral striatum (DLS) accompanies the expression of amphetamine (AMPH) locomotor sensitization. However, whether individual differences in DLS activity previous to AMPH administration can predict the expression of locomotor sensitization has not been assessed. Here, we examined DLS neuronal activity before and after repeated AMPH administration and related it to the susceptibility of rats to sensitize. For that, single-unit recordings on DLS medium spiny neurons (MSNs) were carried out in freely moving male Sprague Dawley rats during repeated AMPH administration. We also examined differences in neurostructure that could accompany sensitization. We quantified the density of the inhibitory postsynaptic marker gephyrin (Geph) in the entopeduncular nucleus (EP) and globus pallidus (GP). A higher burst firing and a lower percentage of correlation between MSNs post-Saline firing rate vs. locomotion predicted the expression of locomotor sensitization. Moreover, during the AMPH challenge, we observed that burst firing decreased in sensitized rats, in contrast to non-sensitized rats in which burst firing was maintained. Finally, a higher Geph density on GP but not EP was observed in non-sensitized rats after AMPH challenge. These results indicate that initial differences in DLS burst firing might underlie the susceptibility to express locomotor sensitization and suggest that the potentiation of dorsal striatum indirect pathway could be considered a protective mechanism to locomotor sensitization.

Tumor necrosis factor-α modulates GABAergic and dopaminergic neurons in the ventrolateral periaqueductal gray of female mice.

Neuroimmune signaling is increasingly identified as a critical component of various illnesses, including chronic pain, substance use disorder, and depression. However, the underlying neural mechanisms remain unclear. Proinflammatory cytokines, such as tumor necrosis factor-α (TNF-α), may play a role by modulating synaptic function and long-term plasticity. The midbrain structure periaqueductal gray (PAG) plays a well-established role in pain processing, and although TNF-α inhibitors have emerged as a therapeutic strategy for pain-related disorders, the impact of TNF-α on PAG neuronal activity has not been thoroughly characterized. Recent studies have identified subpopulations of ventrolateral PAG (vlPAG) with opposing effects on nociception, with dopamine (DA) neurons driving pain relief in contrast to GABA neurons. Therefore, we used slice physiology to examine the impact of TNF-α on neuronal activity of both these subpopulations. We focused on female mice since the PAG is a sexually dimorphic region and most studies use male subjects, limiting our understanding of mechanistic variations in females. We selectively targeted GABA and DA neurons using transgenic reporter lines. Following exposure to TNF-α, there was an increase in excitability of GABA neurons along with a reduction in glutamatergic synaptic transmission. In DA neurons, TNF-α exposure resulted in a robust decrease in excitability along with a modest reduction in glutamatergic synaptic transmission. Interestingly, TNF-α had no effect on inhibitory transmission onto DA neurons. Collectively, these data suggest that TNF-α differentially affects the function of GABA and DA neurons in female mice and enhances our understanding of how TNF-α-mediated signaling modulates vlPAG function.NEW & NOTEWORTHY This study describes the effects of TNF-α on two distinct subpopulations of neurons in the vlPAG. We show that TNF-α alters both neuronal excitability and glutamatergic synaptic transmission on GABA and dopamine neurons within the vlPAG of female mice. This provides critical new information on the role of TNF-α in the potential modulation of pain, since activation of vlPAG GABA neurons drives nociception, whereas activation of dopamine neurons drives analgesia.

Partial protective effects of melatonin on developing brain in a rat model of chorioamnionitis.

Melatonin has shown promising neuroprotective effects due to its anti-oxidant, anti-inflammatory and anti-apoptotic properties, making it a candidate drug for translation to humans in conditions that compromise the developing brain. Our study aimed to explore the impact of prenatal melatonin in an inflammatory/infectious context on GABAergic neurons and on oligodendrocytes (OLs), key cells involved in the encephalopathy of prematurity. An inflammatory/infectious agent (LPS, 300 µg/kg) was injected intraperitoneally (i.p.) to pregnant Wistar rats at gestational day 19 and 20. Melatonin (5 mg/kg) was injected i.p. following the same schedule. Immunostainings focusing on GABAergic neurons, OL lineage and myelination were performed on pup brain sections. Melatonin succeeded in preventing the LPS-induced decrease of GABAergic neurons within the retrospenial cortex, and sustainably promoted GABAergic neurons within the dentate gyrus in the inflammatory/infectious context. However, melatonin did not effectively prevent the LPS-induced alterations on OLs and myelination. Therefore, we demonstrated that melatonin partially prevented the deleterious effects of LPS according to the cell type. The timing of exposure related to the cell maturation stage is likely to be critical to achieve an efficient action of melatonin. Furthermore, it can be speculated that melatonin exerts a modest protective effect on extremely preterm infant brains.

mGlu1 potentiation enhances prelimbic somatostatin interneuron activity to rescue schizophrenia-like physiological and cognitive deficits.

Evidence for prefrontal cortical (PFC) GABAergic dysfunction is one of the most consistent findings in schizophrenia and may contribute to cognitive deficits. Recent studies suggest that the mGlu1 subtype of metabotropic glutamate receptor regulates cortical inhibition; however, understanding the mechanisms through which mGlu1 positive allosteric modulators (PAMs) regulate PFC microcircuit function and cognition is essential for advancing these potential therapeutics toward the clinic. We report a series of electrophysiology, optogenetic, pharmacological magnetic resonance imaging, and animal behavior studies demonstrating that activation of mGlu1 receptors increases inhibitory transmission in the prelimbic PFC by selective excitation of somatostatin-expressing interneurons (SST-INs). An mGlu1 PAM reverses cortical hyperactivity and concomitant cognitive deficits induced by N-methyl-d-aspartate (NMDA) receptor antagonists. Using in vivo optogenetics, we show that prelimbic SST-INs are necessary for mGlu1 PAM efficacy. Collectively, these findings suggest that mGlu1 PAMs could reverse cortical GABAergic deficits and exhibit efficacy in treating cognitive dysfunction in schizophrenia.

5-HT7 receptors enhance inhibitory synaptic input to principal neurons in the mouse basal amygdala.

The basal amygdala (BA) has been implicated in encoding fear and its extinction. The level of serotonin (5-HT) in the BA increases due to arousal and stress related to aversive stimuli. The effects of 5-HT7 receptor (5-HT7R) activation and blockade on the activity of BA neurons have not yet been investigated. In the present study, a transgenic mouse line carrying green fluorescent protein (GFP) reporter gene was used to identify neurons that express the 5-HT7R. GFP immunoreactivity was present mainly in cells that also expressed GAD67 or parvalbumin (PV), the phenotypic markers for GABAergic interneurons. Most cells showing GFP fluorescence demonstrated firing patterns characteristic of BA inhibitory interneurons. Activation of 5-HT7Rs resulted in a depolarization and/or occurrence of spontaneous spiking activity of BA interneurons that was accompanied by an increase in the mean frequency and mean amplitude of spontaneous inhibitory postsynaptic currents (sIPSCs) recorded from BA principal neurons. These effects were blocked by a specific 5-HT7R antagonist, SB269970 and were absent in slices from 5-HT7R knockout mice. Activation of 5-HT7Rs also decreased the mean frequency of spontaneous excitatory postsynaptic currents (sEPSCs) recorded from BA principal neurons, which was blocked by the GABAA receptor antagonist picrotoxin. Neither inhibitory nor excitatory miniature postsynaptic currents (mIPSCs/mEPSCs) were affected by 5-HT7R activation. These results show that in the BA 5-HT7Rs stimulate an activity-dependent enhancement of inhibitory input from local interneurons to BA principal neurons and provide insights about the possible involvement of BA serotonergic receptors in neuronal mechanisms underlying fear memory.

Muscimol Directly Activates the TREK-2 Channel Expressed in GABAergic Neurons through Its N-Terminus.

The two-pore domain K+ (K2P) channel, which is involved in setting the resting membrane potential in neurons, is an essential target for receptor agonists. Activation of the γ-aminobutyric acid (GABA) receptors (GABAAR and GABABR) reduces cellular excitability through Cl- influx and K+ efflux in neurons. Relatively little is known about the link between GABAAR and the K+ channel. The present study was performed to identify the effect of GABAR agonists on K2P channel expression and activity in the neuroblastic B35 cells that maintain glutamic acid decarboxylase (GAD) activity and express GABA. TASK and TREK/TRAAK mRNA were expressed in B35 cells with a high level of TREK-2 and TRAAK. In addition, TREK/TRAAK proteins were detected in the GABAergic neurons obtained from GABA transgenic mice. Furthermore, TREK-2 mRNA and protein expression levels were markedly upregulated in B35 cells by GABAAR and GABABR agonists. In particular, muscimol, a GABAAR agonist, significantly increased TREK-2 expression and activity, but the effect was reduced in the presence of the GABAAR antagonist bicuculine or TREK-2 inhibitor norfluoxetine. In the whole-cell and single-channel patch configurations, muscimol increased TREK-2 activity, but the muscimol effect disappeared in the N-terminal deletion mutant. These results indicate that muscimol directly induces TREK-2 activation through the N-terminus and suggest that muscimol can reduce cellular excitability by activating the TREK-2 channel and by inducing Cl- influx in GABAergic neurons.

Antiparkinsonian-like effects of CPL500036, a novel selective inhibitor of phosphodiesterase 10A, in the unilateral rat model of Parkinson's disease.

Phosphodiesterase 10A (PDE10A), the enzyme which catalyzes hydrolysis of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), is located almost exclusively in striatal γ-amino-butyric acid (GABA)ergic medium spiny neurons (MSNs). Since dopaminergic deficiency in Parkinson's disease (PD) leads to functional imbalance of striatal direct and indirect output pathways formed by MSNs, PDE10A seems to be of special interest as a potential therapeutic target in PD. The aim of the present study was to examine the influence of 7-{5,8-dimethyl-[1,2,4]triazolo[1,5-a]pyrazin-2-yl}-2-phenylimidazo[1,2-a]pyrimidine (CPL500036), a novel selective inhibitor of PDE10A, on sensorimotor deficits and therapeutic effects of L-3,4-dihydroxyphenylalanine (L-DOPA) in hemiparkinsonian rats. Animals were unilaterally lesioned with 6-hydroxydopamine, and their sensorimotor deficits were examined in the stepping, cylinder, vibrissae and catalepsy tests. CPL500036 (0.1 and 0.3 mg/kg) was administered either acutely or chronically (2 weeks), alone or in combination with L-DOPA/benserazide (6 mg/kg/6 mg/kg). Acute treatment with CPL500036 reversed the lesion-induced impairments of contralateral forelimb use in the stepping and cylinder tests but did not influence deficits in the vibrissae test and the lesion-induced catalepsy. Moreover, CPL500036 did not diminish the therapeutic effects produced by acute and chronic treatment with L-DOPA in these tests. The present study suggests a potential use of CPL500036 as a co-treatment to L-DOPA in PD therapy.

Zonisamide attenuates the severity of levodopa-induced dyskinesia via modulation of the striatal serotonergic system in a rat model of Parkinson's disease.

Glutamate, GABA, acetylcholine, dopamine, and serotonin interact with each other to regulate the flow of neural information in the striatum. Serotonin type 1A receptor (5HT1A) is primarily expressed on glutamatergic nerve terminals, and 5HT1B is expressed on GABAergic medium spiny neurons (MSNs). Zonisamide (ZNS) reportedly improves the off period without worsening levodopa-induced dyskinesia (LID) in patients with advanced Parkinson's disease. In this study, LID model rats were prepared by administrating levodopa to unilaterally 6-OHDA-lesioned rats. We analyzed changes in serotonergic neurotransmission of LID model rats to elucidate the relationship between LID and the serotonergic system and pathomechanism of the anti-dyskinetic effects of ZNS. Abnormal involuntary movements (AIMs) were most severe in intermittently levodopa-treated rats but milder in rats intermittently medicated with levodopa and ZNS. Continuously levodopa-infused rats or intermittently ZNS-injected rats did not develop AIMs, and no differences in the expression of brain-derived neurotrophic factor, 5-HT transporter, 5HT1A, and 5HT1B mRNA between the lesioned striatum and normal side were observed. Expression of 5HT1B mRNA was elevated in the lesioned striatum of intermittently levodopa-treated rats, but this elevation was normalized by concomitant use of ZNS. The severity of AIMs was correlated with the ratio of 5HT1B to 5HT1A mRNA expression in the lesioned striatum, indicating that the anti-LID effect of ZNS is based on inhibition via 5HT1B receptors to direct pathway MSNs sensitized by intermittent levodopa treatment. Selectively acting serotonergic drugs, especially those that lower the 5HT1B to 5HT1A ratio, are promising new therapeutic agents to attenuate LID development.

Lateral hypothalamus involvement in control of stress response by bed nucleus of the stria terminalis endocannabinoid neurotransmission in male rats.

The endocannabinoid neurotransmission acting via local CB1 receptor in the bed nucleus of the stria terminalis (BNST) has been implicated in behavioral and physiological responses to emotional stress. However, the neural network related to this control is poorly understood. In this sense, the lateral hypothalamus (LH) is involved in stress responses, and BNST GABAergic neurons densely innervate this hypothalamic nucleus. However, a role of BNST projections to the LH in physiological responses to stress is unknown. Therefore, using male rats, we investigated the role of LH GABAergic neurotransmission in the regulation of cardiovascular responses to stress by CB1 receptors within the BNST. We observed that microinjection of the selective CB1 receptor antagonist AM251 into the BNST decreased the number of Fos-immunoreactive cells within the LH of rats submitted to acute restraint stress. Treatment of the BNST with AM251 also enhanced restraint-evoked tachycardia. Nevertheless, arterial pressure increase and sympathetically-mediated cutaneous vasoconstriction to restraint was not affected by CB1 receptor antagonism within the BNST. The effect of AM251 in the BNST on restraint-evoked tachycardia was abolished in animals pretreated with the selective GABAA receptor antagonist SR95531 in the LH. These results indicate that regulation of cardiovascular responses to stress by CB1 receptors in the BNST is mediated by GABAergic neurotransmission in the LH. Present data also provide evidence of the BNST endocannabinoid neurotransmission as a mechanism involved in LH neuronal activation during stressful events.