Infusion Of Bicuculline Research Articles

The additive effect between citalopram and muscimol upon induction of antinociceptive effect in male mice

Previous investigations have revealed the role of GABAergic and serotonergic systems in the modulation of pain behavior. This research aimed to examine the effects of intracerebroventricular (i.c.v.) infusion of GABAA receptor agonist and antagonist as well as citalopram on pain behavior in male mice. For i.c.v. microinjection, a guide cannula was surgically implanted in the left lateral ventricle of male mice. Pain behavior was evaluated using a tail-flick test. Tail flick latency was measured in each experimental group of mice every 15 min (for 60 min). I.c.v. microinjection of muscimol (0.5 and 1 µg/mouse; GABAA receptor agonist) into the left lateral ventricle dose-dependently induced an antinociceptive effect. On the other hand, i.c.v. infusion of bicuculline (1 µg/mouse; GABAA receptor antagonist) induced a hyperalgesia response. Moreover, intraperitoneally (i.p.) administration of citalopram (8 mg/kg) produced an antinociceptive effect. Co-treatment of citalopram (8 mg/kg) along with muscimol (0.25 µg/mouse) or bicuculline (0.25 µg/mouse) potentiated the antinociceptive effect produced by citalopram. We found an additive antinociceptive effect of citalopram and muscimol in male mice. In conclusion, our results suggested an interaction between citalopram and GABAergic agents on the modulation of pain behavior in male mice.

Chemogenetic attenuation of cortical seizures in nonhuman primates

Epilepsy is a disorder in which abnormal neuronal hyperexcitation causes several types of seizures. Because pharmacological and surgical treatments occasionally interfere with normal brain function, a more focused and on-demand approach is desirable. Here we examined the efficacy of a chemogenetic tool—designer receptors exclusively activated by designer drugs (DREADDs)—for treating focal seizure in a nonhuman primate model. Acute infusion of the GABAA receptor antagonist bicuculline into the forelimb region of unilateral primary motor cortex caused paroxysmal discharges with twitching and stiffening of the contralateral arm, followed by recurrent cortical discharges with hemi- and whole-body clonic seizures in two male macaque monkeys. Expression of an inhibitory DREADD (hM4Di) throughout the seizure focus, and subsequent on-demand administration of a DREADD-selective agonist, rapidly suppressed the wide-spread seizures. These results demonstrate the efficacy of DREADDs for attenuating cortical seizure in a nonhuman primate model.

RF01 | PMON45 Urocortin 3 Interaction with GABA and Glutamate in the Posterodorsal Subnucleus of The Medial Amygdala Mediates Suppression of GnRH Pulse Generator Frequency in Female Mice

Abstract Stress exerts a profound impact on reproductive function and disrupts pulsatile luteinizing hormone (LH) secretion. The posterodorsal subnucleus of the medial amygdala (MePD) is an upstream modulator of the hypothalamus pituitary gonadal axis. Stress alters neuronal activity within the MePD, increasing the expression of Urocortin3 (Ucn3) while enhancing the inhibitory output from the MePD to key hypothalamic reproductive centres. Designer receptor exclusively activated by designer drugs (DREADDs) inhibition of MePD Ucn3 neurons prevents psychological stress-induced suppression of LH pulses in female mice. The neurochemical pathways in the MePD responsible for regulating the GnRH pulse generator in the presence of stress remain unknown. We investigate the neurotransmission involved in the neural circuitry responsible for the suppression of GnRH pulse generator activity by MePD Ucn3 neuronal activation. Ucn3-Cre-tdTomato female ovariectomised (OVX) mice were unilaterally injected with AAV-ChR2 and implanted with optofluid cannulae targeting the MePD. Firstly, we optically stimulated MePD Ucn3 neurons with blue light at 10 Hz, 10 mW and monitored the effect on LH pulses. Next, we combined optogenetic activation of MePD Ucn3 neurons with pharmacological antagonism of GABAA or GABAB receptors with bicuculline or CGP, respectively, and observed the effect on LH pulsatility. Finally, during optical activation of MePD Ucn3 neurons we administered a combination of NMDA and AMPA receptor antagonists, AP5 and CNQX respectively, and monitored the effect on pulsatile LH secretion. Optogenetic stimulation of MePD Ucn3 neurons suppressed pulsatile LH secretion compared to controls and administration of aCSF had no effect on LH pulse interval (MePD Ucn3 activation: 36.04 ± 2.49 vs. pre-treatment control period: 17.50 ± 0.67 vs control virus: 21.67 ± 1.67 min; mean ± SEM; p<0.0001; n=12). Intra-MePD infusion of bicuculline (BIC) or CGP during optogenetic stimulation of MePD Ucn3 neurons completely blocked the suppressive effect of MePD Ucn3 neuronal activation on LH pulsatility (MePD Ucn3 activation and BIC: 20.50 ± 2.03 vs. pre-treatment control period: 17.58 ± 0.75 min; MePD Ucn3 activation and CGP: 16.30 ± 0.63 vs. pre-treatment control period: 17.50 ± 0.77 min; n=7-10). Intra-MePD infusion of AP5 + CNQX during optogenetic stimulation of MePD Ucn3 neurons similarly completely blocked the suppressive effect of MePD Ucn3 neuronal activation on LH pulsatility (MePD Ucn3 activation and AP5 + CNQX: 19.58 ± 1.42 min; n=6). Our findings show for the first time that optogenetic stimulation of MePD Ucn3 neurons inhibits GnRH pulse generator activity via GABA and glutamate signalling within the MePD. Presentation: Saturday, June 11, 2022 1:12 p.m. - 1:17 p.m., Monday, June 13, 2022 12:30 p.m. - 2:30 p.m.

Medial septum activation improves strategy switching once strategies are well-learned via bidirectional regulation of dopamine neuron population activity.

Strategy switching is a form of cognitive flexibility that requires inhibiting a previously successful strategy and switching to a new strategy of a different categorical modality. It is dependent on dopamine (DA) receptor activation and release in ventral striatum and prefrontal cortex, two primary targets of ventral tegmental area (VTA) DA projections. Although the circuitry that underlies strategy switching early in learning has been studied, few studies have examined it after extended discrimination training. This may be important as DA activity and release patterns change across learning, with several studies demonstrating a critical role for substantia nigra pars compacta (SNc) DA activity and release once behaviors are well-learned. We have demonstrated that medial septum (MS) activation simultaneously increased VTA and decreased SNc DA population activity, as well as improved reversal learning via these actions on DA activity. We hypothesized that MS activation would improve strategy switching both early in learning and after extended training through its ability to increase VTA DA population activity and decrease SNc DA population activity, respectively. We chemogenetically activated the MS of male and female rats and measured their performance on an operant-based strategy switching task following 1, 10, or 15 days of discrimination training. Contrary to our hypothesis, MS activation did not affect strategy switching after 1 day of discrimination training. MS activation improved strategy switching after 10 days of training, but only in females. MS activation improved strategy switching in both sexes after 15 days of training. Infusion of bicuculline into the ventral subiculum (vSub) inhibited the MS-mediated decrease in SNc DA population activity and attenuated the improvement in strategy switching. Intra-vSub infusion of scopolamine inhibited the MS-mediated increase in VTA DA population activity but did not affect the improvement in strategy switching. Intra-vSub infusion of both bicuculline and scopolamine inhibited the MS-mediated effects on DA population activity in both the SNc and VTA and completely prevented the improvement in strategy switching. These data indicate that MS activation improves strategy switching once the original strategy has been sufficiently well-learned, and that this may occur via the MS's regulation of DA neuron responsivity.

Optogenetic activation of the reticular nucleus of the thalamus attenuates limbic seizures via inhibition of the midline thalamus.

The nucleus reticularis of the thalamus (nRT) is most studied in epilepsy for its role in the genesis of absence seizures; much less is known regarding its role in other seizure types, including those originating in limbic structures and the temporal lobe. As it is a major source of inhibitory input to higher order thalamic nuclei, stimulation of the nRT may be an effective strategy to disrupt seizure activity that requires thalamic engagement. We recorded single unit activity from the nRT prior to and after infusion of bicuculline into the area tempestas. We monitored single unit activity time-locked with interictal spikes. We optogenetically activated the nRT in both the area tempestas and amygdala kindling models. We tested a role for projections from the nRT to higher order midline thalamic nuclei through the use of retrogradely trafficked viral vector. Mean firing rate in the nRT was decreased after infusion of bicuculline into the area tempestas as compared to the preinfusion baseline. nRT unit firing in response to interictal spikes was heterogeneous, with an approximately equal proportion of neurons displaying (1) no change in firing, (2) increased firing, and (3) decreasing firing. Optogenetic activation of the nRT significantly suppressed seizure activity in both the area tempestas and amygdala kindling models. Optogenetic activation of contralaterally targeting projections but not ipsilaterally targeting projections from the nRT to the midline thalamus significantly suppressed seizures in the kindling model. Although the nRT is typically thought of in the context of absence seizures, our data show that it may be a viable target for other seizure types. In two models that recapitulate the seizure types seen in temporal lobe epilepsy, nRT activation suppressed both electrographic and behavioral seizures. These data suggest that the nRT should be considered more broadly in the context of epilepsy.

Local administration of bicuculline into the ventrolateral and medial preoptic nuclei modifies sleep and maternal behavior in lactating rats.

The preoptic area (POA) is a brain structure classically involved in a wide variety of animal behavior including sleep and maternal care. In the current study, we evaluate the specific effect of disinhibition of two specific regions of the POA, the medial POA nucleus (mPOA) and the ventrolateral POA area (VLPO) on sleep and maternal behavior in lactating rats. For this purpose, mother rats on postpartum day 1 (PPD1) were implanted for polysomnographic recordings and with bilateral cannulae either in the mPOA or in the VLPO. The rats were tested for sleep and maternal behavior on PPD4–8 after the infusion of the GABA-A antagonist, bicuculline (0, 10 or 30 ng/0.2 µl/side). Infusion of bicuculline into the mPOA augmented retrieving and nest building behaviors and reduced both nursing and milk ejections but had almost no effect on sleep. When bicuculine was microinjected into the VLPO, the rats significantly increase the number of retrievings and mouthings and reduced the nursing time without changes in milk ejections, which was associated with an increase in wakefulness and a reduction in light sleep.Our results show that disinhibition of the mPOA, a key area in the control of maternal behavior, increased active maternal behaviors and reduced nursing without affecting wakefulness or sleep time. In contrast, the enhancement of some active maternal behaviors when the drug was infused into the VLPO, a sleep-promoting area, with a concomitant increase in wakefulness suggests that mother rats devote this additional waking time in the active maternal care of the pups. We hypothesize that maternal behavior changes after bicuculine microinjection into the VLPO are caused by a reduction in the sleep drive, rather than a direct effect on maternal behavior.

Generating Acute and Chronic Experimental Models of Motor Tic Expression in Rats.

Motor tics are sudden, rapid, recurrent movements that are the key symptoms of Tourette syndrome and other tic disorders. The pathophysiology of tic generation is associated with abnormal inhibition of the basal ganglia, particularly its primary input structure, the striatum. In animal models of both rodents and non-human primates, local application of GABAA antagonists, such as bicuculline and picrotoxin, into the motor parts of the striatum induces local disinhibition resulting in the expression of motor tics. Here, we present acute and chronic models of motor tics in rats. In the acute model, bicuculline microinjections through a cannula implanted in the dorsal striatum elicit the expression of tics lasting for short time periods of up to an hour. The chronic model is an alternative enabling the extension of tic expression to periods of several days or even weeks, utilizing continuous infusion of bicuculline via a sub-cutaneous mini-osmotic pump. The models enable the study of the behavioral and neural mechanisms of tic generation throughout the cortico-basal ganglia pathway. The models support the implantation of additional recording and stimulation devices in addition to the injection cannulas, thus allowing for a wide variety of usages such as electrical and optical stimulation and electrophysiological recordings. Each method has different advantages and shortcomings: the acute model enables the comparison of the kinematic properties of movement and the corresponding electrophysiological changes before, during and after tic expression and the effects of short-term modulators on tic expression. This acute model is simple to establish; however, it is limited to a short period of time. The chronic model, while more complex, makes feasible the study of tic dynamics and behavioral effects on tic expression over prolonged periods. Thus, the type of empirical query drives the choice between these two complementary models of tic expression.

GABA in the medial prefrontal cortex regulates anxiety-like behavior during the postpartum period

The postpartum period is commonly accompanied by emotional changes, which for many new mothers includes a reduction in anxiety. Previous research in rodents has shown that the postpartum attenuation in anxiety is dependent on offspring contact and has further implicated enhanced GABAergic neurotransmission as an underlying mechanism. However, the specific brain regions where GABA acts to regulate the offspring-induced reduction in postpartum anxiety requires further investigation. Here, we test the hypothesis that offspring interactions suppress anxiety-like behavior in postpartum female rats via GABA signaling in the medial prefrontal cortex (mPFC). Our results show a postpartum reduction in anxiety-like behavior, an effect which was abolished by localized infusion of the GABAA receptor antagonist bicuculline in the mPFC. We also show that activation of GABAA receptors in the mPFC by the agonist muscimol was effective in restoring anxiolyisis in mothers separated from their pups. Lastly, we show that heightened anxiety-like behavior in pup-separated mothers was accompanied by a lower number and percentage of activated GABAergic neurons within the mPFC. Together, these results suggest that mother-offspring interactions reduce anxiety-like behavior in postpartum females via GABAA neurotransmission in the mPFC and in doing so provide insight into mechanisms that may become dysfunctional in mothers who experience high postpartum anxiety.

Central Actions of 3α,5α-THP Involving NMDA and GABAA Receptors Regulate Affective and Sexual Behavior of Female Rats.

The neurosteroid, 5α-pregnan-3α-ol-20-one (known as “allopregnanolone” or 3α,5α-THP), is produced in the midbrain ventral tegmental area (VTA), independent of peripheral sources of progestogens, where it has potential actions at N-methyl-D-aspartate (NMDA) and GABAA receptors to facilitate rodent sexual behavior. Progestogens can also have anti-anxiety effects, but whether these involve actions of centrally-derived 3α,5α-THP or these receptors to support reproductively-relevant behavior is not well understood. We investigated the extent to which 3α,5α-THP’s actions via NMDA and/or GABAA receptors in the midbrain VTA influence reproductive behaviors. Estradiol-primed, ovariectomized/adrenalectomized (OVX/ADX) rats received midbrain VTA infusions of vehicle, an NMDA receptor blocker (MK-801; 200 ng), or a GABAA receptor blocker (bicuculline; 100 ng) followed by a second infusion of vehicle or 3α,5α-THP (100 ng). Reproductively-relevant behaviors were assessed: sexual (paced mating), anxiety-like (elevated plus maze), and social (partner preference, social interaction) behavior. Compared to vehicle, intra-VTA infusions of MK-801 exerted anxiolytic-like effects on elevated plus maze behavior and enhanced lordosis. Unlike prior observations in gonadally-intact rats, intra-VTA bicuculline had no effect on the behavior of OVX/ADX rats (likely due to a floor effect). Subsequent infusions of 3α,5α-THP reversed effects on lordosis and infusions of bicuculline inhibited 3α,5α-THP-facilitated lordosis. Thus, NMDA and GABAA receptors may act as mediators for reproductive behavioral effects of 3α,5α-THP in the midbrain VTA.

Manipulations of inhibition in cortical circuitry differentially affect spectral and temporal features of Bengalese finch song.

The interplay between inhibition and excitation can regulate behavioral expression and control, including the expression of communicative behaviors like birdsong. Computational models postulate varying degrees to which inhibition within vocal motor circuitry influences birdsong, but few studies have tested these models by manipulating inhibition. Here we enhanced and attenuated inhibition in the cortical nucleus HVC (used as proper name) of Bengalese finches (Lonchura striata var. domestica). Enhancement of inhibition (with muscimol) in HVC dose-dependently reduced the amount of song produced. Infusions of higher concentrations of muscimol caused some birds to produce spectrally degraded songs, whereas infusions of lower doses of muscimol led to the production of relatively normal (nondegraded) songs. However, the spectral and temporal structures of these nondegraded songs were significantly different from songs produced under control conditions. In particular, muscimol infusions decreased the frequency and amplitude of syllables, increased various measures of acoustic entropy, and increased the variability of syllable structure. Muscimol also increased sequence durations and the variability of syllable timing and syllable sequencing. Attenuation of inhibition (with bicuculline) in HVC led to changes to song distinct from and often opposite to enhancing inhibition. For example, in contrast to muscimol, bicuculline infusions increased syllable amplitude, frequency, and duration and decreased the variability of acoustic features. However, like muscimol, bicuculline increased the variability of syllable sequencing. These data highlight the importance of inhibition to the production of stereotyped vocalizations and demonstrate that changes to neural dynamics within cortical circuitry can differentially affect spectral and temporal features of song.NEW & NOTEWORTHY We reveal that manipulations of inhibition in the cortical nucleus HVC affect the structure, timing, and sequencing of syllables in Bengalese finch song. Enhancing and blocking inhibition led to opposite changes to the acoustic structure and timing of vocalizations, but both caused similar changes to vocal sequencing. These data provide support for computational models of song control but also motivate refinement of existing models to account for differential effects on syllable structure, timing, and sequencing.

The lateral preoptic area and ventral pallidum embolden behavior

While recently completing a study of the effects of stimulating the lateral preoptic area (LPO) and ventral pallidum (VP) on locomotion and other movements, we also noticed LPO and VP effects on motivational drive and threat tolerance. Here, we have investigated these latter effects by testing conditioned place preference (CPP), behavior on the elevated plus maze (EPM) and the willingness of sated rats to occupy a harshly lit open field center to acquire sweet pellets, a measure of threat tolerance, following infusions of vehicle or bicuculline (bic) into the LPO and VP. LPO-bic infusions robustly increased total locomotion, and, in direct proportion, occupancy of both the harshly lit field center and open arms of the EPM. LPO bic also generated CPP, but did not increase sweet pellet ingestion. These effects were attenuated by dopamine D1 and D2 receptor antagonists, whether given individually or as a cocktail and systemically or infused bilaterally into the nucleus accumbens. VP-bic infusions did not increase total locomotion, but preferentially increased field center occupancy. VP-bic-infused rats compulsively ingested sweet pellets and did so even under the spotlight, whereas harsh illumination suppressed pellet ingestion in the control groups. VP bic produced CPP and increased open arm occupancy on the EPM. These effects were attenuated by pretreatment with dopamine receptor antagonists given systemically or as bilateral infusions into the VP, except for % distance in the field center (by D1 or D2 antagonists) and pellet ingestion (by D1 antagonist). Thus, boldness generated in association with LPO activation is tightly tied to locomotor activation and, as is locomotion itself, strongly DA dependent, whereas that accompanying stimulation of the VP is independent of locomotor activation and, at least in part, DA signaling. Furthermore, respective emboldened behaviors elicited from neither LPO nor VP could clearly be attributed to goal pursuit. Rather, emboldening of behavior seems more to be a fixed action response not fundamentally different than previously for reported locomotion, pivoting, backing, gnawing, and eating elicited by basal forebrain stimulation.

Medial septum activation produces opposite effects on dopamine neuron activity in the ventral tegmental area and substantia nigra in MAM vs. normal rats

The medial septum (MS) differentially impacts midbrain dopamine (DA) neuron activity via the ventral hippocampus, a region implicated in DA-related disorders. However, whether MS regulation of ventral tegmental area (VTA) and substantia nigra pars compacta (SNc) is disrupted in a developmental disruption model of schizophrenia is unknown. Male Sprague-Dawley rats were exposed at gestational day 17 to methylazoxymethanol (MAM) or saline. As adults, NMDA (0.75 µg/0.2 µL) was infused into the MS, and either DA neuron activity in the VTA and SNc (7–9 anesthetized rats per group) or amphetamine-induced hyperlocomotion (AIH, 11-13 rats per group) was measured. MS activation produced a 58% increase in the number of spontaneously active DA neurons in VTA and a 37% decrease in SNc in saline rats. However, MS activation produced opposite effects on DA population activity in MAM rats, decreasing VTA DA activity by 51% and increasing SNc DA activity by 47%. MS activation also increased AIH by 113% in MAM rats, opposite of what is seen in intact rats. The effect in behavioral output may be due to disrupted GABAergic regulation of SNc as bicuculline infusion into vSub, which selectively prevented the MS activation-induced decrease in SNc DA activity in intact rats, prevented the increase in AIH and SNc DA activity in MAM rats. These findings demonstrate that the regulation of midbrain DA neurons by the MS is disrupted in this well-validated animal model, suggesting that it could be a potential locus for pharmacological intervention in disorders such as schizophrenia.

Deciphering a GABAergic Neural Circuit in Critical Control of Leptin-Mediated Feeding, Body Weight, and Glucose Homeostasis

Leptin plays a major role in control of glucose metabolism and feeding behavior by acting upon the LepR neurons that widely distribute through the brain. Although several LepR-expressing neuronal groups were identified in distinct nuclei, a dominant neural circuit that mediates the physiological roles of leptin remains elusive, largely due to the inadequate phenotypes as a result of developmental and neural compensations. In this report, a novel, non-sense suppression-based, inducible genetic system is implemented to rapidly delete a specific group of LepR signaling from adult mice within 4 days. Acute deletion of LepR from AgRP neurons show a drastic increase of food intake, massive weight gain, and severe glucose intolerance, developing in a rate with tight manifestation of the global leptin deficiency. Furthermore, a neural circuit comprising of LepR-expressing AgRP neurons and downstream GABAergic neurons in the DMH is mapped by genetic and functional tracing approaches. Optogenetic manipulation of AgRPLepR-DMHGABA neural circuit or the post-synaptic DMHMC4R neurons strongly altered food intake and glucose tolerance in a bidirectional controlled manner. Among all GABAA subunits examined, Gabra3 (encoding GABAA α3 subunits) displayed the most robust changes in response to genetic inactivation of LepRAgRP signaling. Using CRISPR/Cas9 gene editing technique, loss of Gabra3 in DMHMC4R neurons reduced food intake and enhanced glucose intolerance. Surprisingly, chronic infusion of bicuculline into the DMH fully reversed the obesity and glucose intolerance as a result of acute deletion of LepRAgRP signaling. Furthermore, genetic ablation of GABADMH neurons blunted leptin-mediated control of glucose metabolism and food intake. In conclusion, we unveil a novel AgRPLepR-DMHGABA neural circuit and associated GABAA receptor signaling system in fundamental control of leptin-associated obesity and diabetes. Disclosure Q. Wu: None. Y. Han: None.

Lateral preoptic and ventral pallidal roles in locomotion and other movements.

The lateral preoptic area (LPO) and ventral pallidum (VP) are structurally and functionally distinct territories in the subcommissural basal forebrain. It was recently shown that unilateral infusion of the GABAA receptor antagonist, bicuculline, into the LPO strongly invigorates exploratory locomotion, whereas bicuculline infused unilaterally into the VP has a negligible locomotor effect, but when infused bilaterally, produces vigorous, abnormal pivoting and gnawing movements and compulsive ingestion. This study was done to further characterize these responses. We observed that bilateral LPO infusions of bicuculline activate exploratory locomotion only slightly more potently than unilateral infusions and that unilateral and bilateral LPO injections of the GABAA receptor agonist muscimol potently suppress basal locomotion, but only modestly inhibit locomotion invigorated by amphetamine. In contrast, unilateral infusions of muscimol into the VP affect basal and amphetamine-elicited locomotion negligibly, but bilateral VP muscimol infusions profoundly suppress both. Locomotor activation elicited from the LPO by bicuculline was inhibited modestly and profoundly by blockade of dopamine D2 and D1 receptors, respectively, but was not entirely abolished even under combined blockade of dopamine D1 and D2 receptors. That is, infusing the LPO with bic caused instances of near normal, even if sporadic, invigoration of locomotion in the presence of saturating dopamine receptor blockade, indicating that LPO can stimulate locomotion in the absence of dopamine signaling. Pivoting following bilateral VP bicuculline infusions was unaffected by dopamine D2 receptor blockade, but was completely suppressed by D1 receptor blockade. The present results are discussed in a context of neuroanatomical and functional organization underlying exploratory locomotion and adaptive movements.

S40. MEDIAL SEPTUM ACTIVATION PRODUCES OPPOSITE EFFECTS ON DOPAMINE NEURON ACTIVITY IN THE VENTRAL TEGMENTAL AREA AND SUBSTANTIA NIGRA PARS COMPACTA IN MAM VERSUS NORMAL RATS

BackgroundDisruptions in dopamine (DA) signaling are central to the pathophysiology of several major psychiatric disorders, including schizophrenia. Thus, discovery of novel therapeutic approaches that normalize DA signaling is a major focus of research. One pathway that is critical for DA regulation originates from the ventral subiculum (vSub) of the hippocampus and controls ventral tegmental area (VTA) DA neuron activity. A potent regulator of hippocampal function is the medial septum (MS); a sub-region of the basal forebrain that widely innervates the hippocampus, including the vSub, via cholinergic, GABAergic, and glutamatergic projections, drives hippocampal theta rhythms, and affects goal-directed learning and memory. Despite this, it has never been determined if the MS is an afferent regulator of the midbrain DA system, and therefore may be a novel therapeutic treatment target for DA-related disorders.MethodsEffects of MS activation (NMDA, 0.75 µg/ 0.2 µL) were examined in intact and methylazoxymethanol acetate- treated (MAM) male Sprague-Dawley rats using anesthetized single unit DA recordings in the VTA and substantia nigra pars compacta (SNc) and locomotor behavior in an open field following systemic amphetamine (0.75 mg/kg).ResultsMS activation produced a prolonged 71% increase in the number of spontaneously active DA cells in the VTA, and an opposing 40% decrease in the number of active DA cells in the SNc, compared to vehicle infusions. These effects were mediated by the vSub and ventral pallidum as local infusion of TTX (1 µM /0.5 µL) and bicuculline (1 ng/ 0.5 µL), respectively, reversed DA population activity changes in both regions. MS activation also decreased the locomotor response to amphetamine (49% reduction in distance traveled during peak ambulation compared to vehicle). MS activation-induced changes in both DA population activity and amphetamine-induced hyperlocomotion were selectively mediated by different neurotransmitter populations from MS to vSub as infusion of scopolamine (8 µg/1.0 µL) into the vSub selectively prevented DA population activity changes in the VTA. In contrast, infusion of bicuculline (12.5 ng/0.5 µL) selectively prevented DA population activity changes in the SNc and the decrease in amphetamine-induced hyperlocomotion. In MAM rats, MS activation produced opposite effects on DA population activity versus controls as it decreased VTA DA activity by 51% and increased SNc DA activity by 47%. This was accompanied by a similar reversal in amphetamine-induced hyperlocomotion, with MS activation increasing locomotion in MAM animals (113% increase in distance traveled during peak ambulation compared to control animals). The reversal in behavioral output is likely due to disrupted GABAergic projections from MS to vSub as bicuculline infusion into vSub prevented the increase in locomotor behavior.DiscussionThese data indicate that the MS differentially regulates both VTA and SNc DA neuron activity and behavioral output, but via distinct pathways, and that this regulation is disrupted in a well-validated animal model of schizophrenia. Therefore, this suggests that in normal animals the MS might activate the VTA to increase information processing while delaying action via SNc inhibition. In contrast, the inhibition of VTA and activation of SNc in MAM rats might promote a rapid response without adequate processing of information.

Lateral septum stimulation disinhibits dopaminergic neurons in the antero-ventral region of the ventral tegmental area: Role of GABA-A alpha 1 receptors

The mechanisms commanding the activity of dopaminergic neurons of the ventral tegmental area (VTA) and the location of these neurons are relevant for the coding and expression of motivated behavior associated to reward-related signals. Anatomical evidence shows that several brain regions modulate VTA dopaminergic neurons activity via multiple mechanisms. However, there is still scarce knowledge of how the lateral septum (LS) modulates VTA activity. We performed in-vivo dual-probe microdialysis to measure VTA dopamine, glutamate and GABA extracellular levels after LS stimulation in the presence or absence of GABAergic antagonists. Anterograde tracing and immunohistochemical analysis was used to reveal the anatomical relationship between LS and VTA. LS stimulation significantly increased dopamine and GABA, but not glutamate, VTA extracellular levels. Intra VTA infusion of bicuculline, GABA-A receptor antagonist, inhibited the increase of dopamine but not of GABA VTA levels induced by LS stimulation. Intra VTA infusion of indiplon, selective positive allosteric modulator of GABA-A receptors containing alpha1 subunit, significantly increases VTA dopamine extracellular levels induced by LS. Combined c-Fos and tyrosine hydroxylase immunohistochemistry, revealed that LS stimulation increases the activity of dopaminergic neurons in the antero-ventral region of the VTA. Consistently, anterograde tracing with biotinylated dextran amine revealed the existence of fibers arising from the LS to the antero-ventral region of the VTA. Taken together, our results suggest that LS modulates dopaminergic activity in the antero-ventral region of VTA by inhibiting GABAergic interneurons bearing GABA-A receptors containing alpha1 subunit.

Effects of chemical stimulation of the lateral wings of the dorsal raphe nucleus on panic-like defensive behaviors and Fos protein expression in rats

The lateral wings subnucleus of the dorsal raphe nucleus (lwDR) has been implicated in the modulation of panic-like behaviors, such as escape. Infusion of non- excitotoxic doses of the excitatory amino acid kainic acid into this subnucleus promptly evokes a vigorous escape response. In addition, rats exposed to panic-inducing situations show an increase in Fos protein expression in neurons within the lwDR. In the present study, we first investigated whether key structures associated with the mediation of escape behavior are recruited after chemical stimulation of the lwDR with kainic acid. We next investigated whether the infusion of the GABAA receptor antagonist bicuculline into the lwDR also evoked escape responses measured both in a circular arena and in the rat elevated T-maze. The effects of bicuculline in the circular arena were compared to those caused by the infusion of this antagonist into the ventrolateral periaqueductal gray (vlPAG), an area in close vicinity to the lwDR. The results showed that kainic acid infusion into the lwDR increased Fos protein immunostaining in brain structures deeply involved in panic-like defensive behaviors, such as the periaqueductal gray and hypothalamus, but not the amygdala. As observed with kainic acid, bicuculline evoked a pronounced escape response in the circular arena when microinjected in the lwDR, but not in the vlPAG. The escape-promoting effect of bicuculline in the lwDR was also evidenced in the elevated T-maze. These findings strength the view that dysfunction in mechanisms controlling escape in the lwDR is critically implicated in the pathophysiology of panic disorder.

Prolonged striatal disinhibition as a chronic animal model of tic disorders

BackgroundExperimental findings and theoretical models have associated Tourette syndrome with abnormal striatal inhibition. The expression of tics, the hallmark symptom of this disorder, has been transiently induced in non-human primates and rodents by the injection of GABAA antagonists into the striatum, leading to temporary disinhibition. New methodThe novel chronic model of tic expression utilizes mini-osmotic pumps implanted subcutaneously in the rat’s back for prolonged infusion of bicuculline into the dorsolateral striatum. ResultsTics were expressed on the contralateral side to the infusion over a period of multiple days. Tic expression was stable, and maintained similar properties throughout the infusion period. Electrophysiological recordings revealed the existence of tic-related local field potential spikes and individual neuron activity changes that remained stable throughout the infusion period. Comparison with existing methodsThe striatal disinhibition model provides a unique combination of face validity (tic expression) and construct validity (abnormal striatal inhibition) but is limited to sub-hour periods. The new chronic model extends the period of tic expression to multiple days and thus enables the study of tic dynamics and the effects of behavior and pharmacological agents on tic expression. ConclusionsThe chronic model provides similar behavioral and neuronal correlates of tics as the acute striatal disinhibition model but over prolonged periods of time, thus providing a unique, basal ganglia initiated model of tic expression. Chronic expression of symptoms is the key to studying the time varying properties of Tourette syndrome and the effects of multiple internal and external factors on this disorder.

Chronic GABAergic blockade in the spinal cord in vivo induces motor alterations and neurodegeneration

Inhibitory GABAergic and glycinergic neurotransmission in the spinal cord play a central role in the regulation of neuronal excitability, by maintaining a balance with the glutamate-mediated excitatory transmission. Glutamatergic agonists infusion in the spinal cord induce motor neuron death by excitotoxicity, leading to motor deficits and paralysis, but little is known on the effect of the blockade of inhibitory transmission. In this work we studied the effects of GABAergic and glycinergic blockade, by means of microdialysis perfusion (acute administration) and osmotic minipumps infusion (chronic administration) of GABA and glycine receptors antagonists directly in the lumbar spinal cord. We show that acute glycinergic blockade with strychnine or GABAergic blockade with bicuculline had no significant effects on motor activity and on motor neuron survival. However, chronic bicuculline infusion, but not strychnine, induced ipsilateral gait alterations, phalange flaccidity and significant motor neuron loss, and these effects were prevented by AMPA receptor blockade with CNQX but not by NMDA receptor blockade with MK801. In addition, we demonstrate that the chronic infusion of bicuculline enhanced the excitotoxic effect of AMPA, causing faster bilateral paralysis and increasing motor neuron loss. These findings indicate a relevant role of GABAergic inhibitory circuits in the regulation of motor neuron excitability and suggest that their alterations may be involved in the neurodegeneration processes characteristic of motor neuron diseases such as amyotrophic lateral sclerosis.

GABAergic effect on resting-state functional connectivity: Dynamics under pharmacological antagonism

Resting state functional connectivity MRI measures synchronous activity among brain regions although the mechanisms governing the temporally coherent BOLD signals remain unclear. Recent studies suggest that γ-amino butyric acid (GABA) levels are correlated with functional connectivity. To understand whether changes in GABA transmission alter functional connectivity, we modulated the GABAergic activity by a GABAA receptor antagonist, bicuculline. Resting and evoked electrophysiology and BOLD signals were measured in isoflurane-anesthetized rats under infusion of low-dose bicuculline or vehicle individually. Both somatosensory BOLD activations and evoked potentials induced by forepaw stimulation were increased significantly under bicuculline compared to vehicle, indicating increased excitability. Gradually elevated resting BOLD correlation within and between the somatosensory and visual cortices, as well as between somatosensory and caudate putamen but not within subcortical areas were found with the infusion of bicuculline. Increased cerebral blood flow was observed throughout the cortical and subcortical areas where the receptor density is high, but it didn’t correlate with BOLD connectivity except in the primary somatosensory cortex. Furthermore, resting EEG coherence in the alpha and beta bands exhibited consistent change with the BOLD correlation. The increased cortico-cortical and cortico-striatal connectivity without dependence on the receptor distribution indicate that the functional connectivity may be mediated by long-range projection via the cortical and striatal GABAergic inter-neurons. Our results indicate an important role of the GABAergic system on neural and hemodynamic oscillations, which further supports the neuronal basis of functional connectivity MRI and its correlation with neurotransmission.