Cholinergic modulation of purinergic and GABAergic co-transmission at in vitro hypothalamic synapses

The lateral hypothalamus (LH) is an important center for the integration of autonomic and limbic information and is implicated in the modulation of visceral motor and sensory pathways, including those underlying feeding and arousal behaviors. LH neurons in vitro release both ATP and GABA. The control of ATP and GABA co-transmission in LH may underlie the participation of LH in basic aspects of arousal and reinforcement. LH neurons receive cholinergic input from the pedunculopontine and laterodorsal tegmental nuclei as well as from cholinergic interneurons within the LH per se. This study presents evidence for nicotinic acetylcholine receptor (nAChR)-mediated enhancement of GABAergic, but not of purinergic, transmission despite the co-transmission of ATP and GABA at LH synapses in vitro. Facilitation of GABAergic transmission by nicotine is inhibited by antagonists of (alphabeta)*-containing nAChRs, but is unaffected by an alpha7-selective antagonist, consistent with a nAChR-mediated enhancement of GABA release mediated by non-alpha7-containing nAChRs. Activation of muscarinic ACh receptors enhances the release of ATP while concomitantly depressing GABAergic transmission. The independent modulation of ATP/GABAergic transmission may provide a new level of synaptic flexibility in which individual neurons utilize more than one neurotransmitter but retain independent control over their synaptic activity.     

Kainate receptor (GluR5)-mediated disinhibition of responses in rat ventrobasal thalamus allows a novel sensory processing mechanism

Kainate receptors have been studied extensively in vitro , but how they might function physiologically remains unclear. We studied kainate receptor modulation of synaptic responses in the rat ventrobasal thalamus using the novel antagonist LY382884 and the agonist ATPA (selective for GluR5-containing kainate receptors) as tools. No evidence could be found for a direct contribution of kainate receptors to responses of thalamic relay cells to lemniscal (sensory) input in thalamic slices studied with the aid of intracellular and field potential recordings, using selective AMPA and NMDA receptor antagonists and LY382884. However, the GluR5 agonist ATPA reduced the IPSPs originating from the thalamic reticular nucleus. Extracellular single-neurone recordings in anaesthetised rats showed that excitatory responses evoked by physiological vibrissa afferent stimulation were reduced by LY382884 applied iontophoretically at the recording site. This action of the antagonist was occluded when GABA receptors were blocked, indicating that the reduction in excitatory sensory responses by LY382884 is due to an action on GABAergic inhibition arising from the thalamic reticular nucleus. Further experiments showed that these actions depended on whether inhibition was evoked during activation of the excitatory receptive field rather than when inhibition was evoked from a surround vibrissa. We suggest that GluR5 is located presynaptically on inhibitory GABAergic terminals of thalamic reticular nucleus neurones, and that it is normally activated by glutamate spillover from synapses between excitatory afferents and relay neurones during physiological stimulation. We propose that this GluR5-activated disinhibition has an important novel role in extracting sensory information from background noise.     

Activation of D1 dopamine receptors stimulates the release of GABA in the basal ganglia of the rat

Here we have explored whether dopamine is able to modulate the release of gamma-aminobutyric acid (GABA) from striatal terminals to substantia nigra pars reticulata, entopeduncular nucleus, globus pallidus and caudate-putamen. The type of dopamine receptors involved was assessed by the blocking effect of either SCH 23390 (D1 antagonist) or (-)-sulpiride (D2 antagonist) of the dopamine effect. Dopamine stimulated (EC50 3.2 microM) the depolarization-induced release of [3H]GABA from slices isolated from all of the above mentioned nuclei. SCH 23390 dose-dependently blocked the dopamine stimulation, but (-)-sulpiride did not show any blocking effect. The results suggest that dopamine via D1 receptors modulates the release of GABA from striatal GABAergic terminals.     

Intracranial self-stimulation induces Fos expression in GABAergic neurons in the rat mesopontine tegmentum

The cholinergic neurons which originate in the mesopontine tegmentum and innervate the midbrain ventral tegmental area have been proposed to play a key role in intracranial self-stimulation reward. This mesopontine area also contains GABA neurons. Detailed information is still lacking, however, about the relationship of cholinergic and GABAergic neurons in this region to self-stimulation reward. Therefore, using double immunostaining for Fos as a marker of neuronal activity and choline acetyltransferase as a marker of cholinergic neurons, or for Fos and GABA, we investigated whether self-stimulation of the medial forebrain bundle induces Fos expression within cholinergic and GABAergic neurons in two regions of the mesopontine tegmentum, i.e., pedunculopontine tegmental nucleus and laterodorsal tegmental nucleus. Self-stimulation of the medial forebrain bundle for 1 h induced a large increase in the number of cells expressing Fos in both the pedunculopontine tegmental nucleus and laterodorsal tegmental nucleus, when compared to control brains. However, the self-stimulation-induced expression of Fos was restricted mostly to GABA-, but not choline acetyltransferase-, immunostained cells. We also examined, using microdialysis, whether self-stimulation increases acetylcholine efflux in the ventral tegmental area, a terminal region of the mesopontine tegmentum cholinergic pathway. One hour of self-stimulation significantly increased acetylcholine efflux from this terminal area.These results indicate that intracranial self-stimulation of the medial forebrain bundle may increase acetylcholine release without affecting expression of Fos in cholinergic neurons, while the same stimulation may induce Fos expression in GABAergic neurons of the mesopontine tegmentum. GABAergic as well as cholinergic neurons in this area appear to be activated by self-stimulation reward in the medial forebrain bundle.     

GABAergic actions on cholinergic laterodorsal tegmental neurons: implications for control of behavioral state

Cholinergic neurons of the pontine laterodorsal tegmentum (LDT) play a critical role in regulation of behavioral state. Therefore, elucidation of mechanisms that control their activity is vital for understanding of how switching between wakefulness, sleep and anesthetic states is effectuated. In vivo studies suggest that GABAergic mechanisms within the pons play a critical role in behavioral state switching. However, the postsynaptic, electrophysiological actions of GABA on LDT neurons, as well as the identity of GABA receptors present in the LDT mediating these actions is virtually unexplored. Therefore, we studied the actions of GABA agonists and antagonists on cholinergic LDT cells by performing patch clamp recordings in mouse brain slices. Under conditions where detection of Cl(-) -mediated events was optimized, GABA induced gabazine (GZ)-sensitive inward currents in the majority of LDT neurons. Post-synaptic location of GABA(A) receptors was demonstrated by persistence of muscimol-induced inward currents in TTX and low Ca(2+) solutions. THIP, a selective GABA(A) receptor agonist with a preference for δ-subunit containing GABA(A) receptors, induced inward currents, suggesting the existence of extrasynaptic GABA(A) receptors. LDT cells also possess GABA(B) receptors as baclofen-activated a TTX- and low Ca(2+)-resistant outward current that was attenuated by the GABA(B) antagonists CGP 55845 and saclofen. The tertiapin sensitivity of baclofen-induced outward currents suggests that a G(IRK) mediated this effect. Further, outward currents were never additive with those induced by application of carbachol, suggesting that they were mediated by activation of GABA(B) receptors linked to the same G(IRK) activated in these cells by muscarinic receptor stimulation. Activation of GABA(B) receptors inhibited Ca(2+) increases induced by a depolarizing voltage step shown previously to activate VOCCs in cholinergic LDT neurons. Baclofen-mediated reductions in depolarization-induced Ca(2+) were unaltered by prior emptying of intracellular Ca(2+) stores, but were abolished by low extracellular Ca(2+) and pre-application of nifedipine, indicating that activation of GABA(B) receptors inhibits influx of Ca(2+) involving L-type Ca(2+) channels. Presence of GABA(C) receptors is suggested by the induction of inward current by (E)-4- amino-2-butenoic acid (TACA) and its inhibition by 1,2,5,6-tetrahydropyridine-4-ylmethylphosphinic (TPMPA), a relatively selective agonist and antagonist, respectively, of GABA(C) receptors. All of these GABA-mediated actions were found to occur in histochemically-identified cholinergic neurons. Taken together, these data indicate for the first time that cholinergic neurons of the LDT exhibit functional GABA(A, B and C) receptors, including extrasynaptically located GABA(A) receptors, which may be tonically activated by synaptic overflow of GABA. Accordingly, the activity of cholinergic LDT neurons is likely to be significantly affected by GABAergic tone within the nucleus, and so, demonstrated effects of GABA on behavioral state may be mediated, in part, via direct actions on cholinergic neurons in the LDT.     

Evidence for cholinergic muscarinic receptors on mediodorsal thalamic projections to the anterior cingulate cortex

Cholinergic muscarinic receptor binding was analyzed in the rat brain anterior cingulate cortex following lesions of the mediodorsal or anterior thalamic nuclei, or the diagonal band of Broca. A significant change in receptor binding was noted only after lesions of the mediodorsal projection, suggesting that cholinergic muscarinic receptors are located on these terminals. These findings suggest that the projection from the diagonal band of Broca which is cholinergic may act as a modulator of the mediodorsal thalamic projection.     

Muscarinic receptors mediate enhancement of spontaneous GABA release in the chick brain

The functional role of muscarinic acetylcholine receptors in the lateral spiriform nucleus was studied in chick brain slices. Whole-cell patch-clamp recordings of neurons in the lateral spiriform nucleus revealed that carbachol enhanced GABAergic spontaneous inhibitory postsynaptic currents. The duration of the response to carbachol was significantly reduced after blockade of muscarinic receptors with atropine. In the presence of the nicotinic receptor antagonist dihydro-beta-erythroidine, carbachol produced a delayed but prolonged enhancement of spontaneous GABAergic inhibitory postsynaptic currents that was completely blocked by atropine. Muscarine also enhanced the frequency of spontaneous GABAergic inhibitory postsynaptic currents in a dose-dependent manner, but had no effect on inhibitory postsynaptic current amplitude. While 4-diphenylacetoxy-N-(2-chloroethyl)-piperidine hydrochloride, a M3 antagonist, completely blocked muscarine's effect, telenzepine, a M1 antagonist, and tropicamide, a M4 antagonist, only partially decreased the response to muscarine. Pirenzepine, a M1 antagonist, and methoctramine, a M2 antagonist, potentiated muscarine's enhancement of spontaneous GABAergic inhibitory postsynaptic currents. Muscarine's action was blocked by tetrodotoxin, cadmium chloride and omega-conotoxin GVIA, but was not affected by dihydro-beta-erythroidine, 6-cyano-7-nitroquinoxaline-2,3-dione, D(-)-2-amino-5-phosphonopentanoic acid, naloxone or fluphenazine. These results demonstrate that activation of both muscarinic and nicotinic acetylcholine receptors can enhance GABAergic inhibitory postsynaptic currents in the lateral spiriform nucleus. The muscarinic response has a slower onset but lasts longer than the nicotinic effect. The M3 receptor subtype is predominantly involved in enhancing spontaneous GABAergic inhibitory postsynaptic currents. These M3 receptors must be located some distance from GABA release sites, since activation of voltage-dependent sodium channels, and consequent activation of N-type voltage-dependent calcium channels, is required to trigger enhanced GABA release following activation of muscarinic receptors.     

Characterization of the extent of pontomesencephalic cholinergic neurons' projections to the thalamus: comparison with projections to midbrain dopaminergic groups

We sought to determine whether pontomesencephalic cholinergic neurons which we have been shown previously to project to the substantia nigra and ventral tegmental area also contribute to the thalamic activation projection from the pedunculopontine and laterodorsal tegmental nuclei. Retrograde tracing, immunohistochemical localization of choline acetyltransferase and statistical methods were used to determine the full extent of the cholinergic projection from the pedunculopontine and laterodorsal tegmental nuclei to the thalamus. Progressively larger Fluoro-Gold injections in to the thalamus proportionally labeled increasing numbers of pontomesencephalic cholinergic cells both ipsi- and contralaterally in the pedunculopontine and laterodorsal tegmental nuclei. Multiple large thalamic injections left only a small fraction of the ipsilateral pontomesencephalic cholinergic group unlabeled. This small remainder did not correspond to the populations which project to the substantia nigra and ventral tegmental area, thereby indicating that substantia nigra- and ventral tegmental area-projecting cholinergic neurons must also project to the thalamus. We examined whether there existed any set of cholinergic neurons in the pedunculopontine and laterodorsal tegmental nuclei which did not innervate a thalamic target. The distribution of descending projections of the pedunculopontine and laterodorsal tegmental nuclei demonstrated that the unlabeled remainder cannot correspond to a purely descending group. We also show that substance P-positive cholinergic cells in the laterodorsal tegmental nucleus project to the thalamus. Further studies demonstrated that the small population of cholinergic cells left unlabeled from the thalamus were the smallest sized cholinergic cells, and included two groups of small, light-staining cholinergic cells located in the parabrachial area and central gray, adjacent to the main pedunculopontine and laterodorsal tegmental nuclei cholinergic groups. These small cells, in contrast to thalamic-projecting cholinergic cells, did not stain positively for reduced nicotinamide adenine dinucleotide phosphate-diaphorase. Taken together, these results indicated that all of the reduced nicotinamide adenine dinucleotide phosphate diaphorase-positive/choline acetyltransferase-positive neurons of the pedunculopontine/laterodorsal tegmental nuclei ascend to innervate some portion of the thalamus, in addition to the other targets they innervate. These findings indicate that the diverse physiological and behavioral effects attributed to the activity of pontomesencephalic cholinergic neurons should not be dissociated from their activating effects in the thalamus.     

Retrograde signaling changes the venue of postsynaptic inhibition in rat substantia nigra

Both endocannabinoids through cannabinoid receptor type I (CB1) receptors and dopamine through dopamine receptor type D1 receptors modulate postsynaptic inhibition in substantia nigra by changing GABA release from striatonigral terminals. By recording from visually identified pars compacta and pars reticulata neurons we searched for a possible co-release and interaction of endocannabinoids and dopamine. Depolarization of a neuron in pars reticulata or in pars compacta transiently suppressed evoked synaptic currents which were blocked by GABA(A) receptor antagonists (inhibitory postsynaptic currents [IPSCs]). This depolarization-induced suppression of inhibition (DSI) was abrogated by the cannabinoid CB1 receptor antagonist AM251 (1 microM). A correlation existed between the degree of DSI and the degree of reduction of evoked IPSCs by the CB1 receptor agonist WIN55,212-2 (1 microM). The cholinergic receptor agonist carbachol (0.5-5 microM) enhanced DSI, but suppression of spontaneous IPSCs was barely detectable pointing to the existence of GABA release sites without CB1 receptors. In dopamine, but not in GABAergic neurons DSI was enhanced by the dopamine D1 receptor antagonist SCH23390 (3-10 microM). Both the antagonist for CB1 receptors and the antagonist for dopamine D1 receptors enhanced or reduced, respectively, the amplitudes of evoked IPSCs. This tonic influence persisted if the receptor for the other ligand was blocked. We conclude that endocannabinoids and dopamine can be co-released. Retrograde signaling through endocannabinoids and dopamine changes inhibition independently from each other. Activation of dopamine D1 receptors emphasizes extrinsic inhibition and activation of CB1 receptors promotes intrinsic inhibition.     

The dopamine-γ aminobutyric acid interaction in the striatum of the rat is differently regulated by dopamine D-1 and D-2 types of receptor: evidence obtained with rotational behavioural experiments

The effects of GABA antagonists on apomorphine-and pergolide-induced rotational behaviour were studied with models combining intracerebral and systemic pharmacological treatments. Whether given systemically or intrastriatally to 6-hydroxydopaminelesioned rats, the GABA antagonist picrotoxin inhibited the rotational responses produced by s. c. administration of the dopamine (DA) D-1/D-2 agonist apomorphine while it enhanced the rotational behaviour produced by the DA D-2 agonist pergolide. Following unilateral injection of picrotoxin or bicuculline into the striatum of naive rats, apomorphine produced ispsilateral rotation, while pergolide produced contralateral rotation. These contrasting effects are compared to the behavioural responses produced by intracerebral administration of GABAergic drugs alone. Intrapallidal injection of picrotoxin produced contralateral rotational behaviour which was independent of pallido-nigral pathways. Contralateral rotation was also produced by GABA agonists, but only following intranigral injections. The results are discussed in terms of differences in the localization of DA D-1 and DA D-2 receptors on striatal GABAergic neurons. The DA D-2 receptor agonist pergolide may induce inhibition of striato-pallidal GABAergic neurons, as well as of a local GABAergic circuit exerting inhibition on a striato-pallidal enkephalinergic pathway. However, the DA D-1/D-2 receptor agonist apomorphine may inhibit striatal interneurons exerting inhibition on a striato-nigral GABAergic projection. Such a neuronal arrangement may explain that striatal DA stimulation increases GABA release from the striato-nigral terminals.     

Presynaptic muscarinic receptors in guinea pig superior cervical ganglion

This study characterizes the presynaptic muscarinic cholinergic receptors associated with the modulation of the electrically-evoked acetylcholine output from guinea pig superior cervical ganglion preincubated with [3H]choline. The M1-selective agonist pilocarpine had no effect while carbachol and oxotremorine strongly decreased the evoked outflow of tritium. Atropine increased such evoked release of [3H]acetylcholine whereas the M1-selective antagonist pirenzepine was ineffective. Moreover, atropine but not pirenzepine antagonized the inhibitory effect of carbachol. These results suggest that the guinea-pig superior cervical ganglion is equipped with presynaptic inhibitory muscarinic receptors of the M2 subtype.     

The role of the laterodorsal tegmental nucleus of the rat in experimental seizures

This study determined the effects of discrete microinjections of GABA agonists in the cholinergic nuclei of the pontomesencephalic tegmentum on spontaneous behavior and seizures induced by intravenous pentylenetetrazol, bicuculline or strychnine, in the rat. Injections of both the GABAA agonist piperidine-4-sulfonic acid and the GABAB agonist (-)baclofen in the laterodorsal tegmental nucleus produced a dose-dependent suppression of behavioral arousal and a reduction in the threshold of myoclonic and clonic but not tonic seizures induced by bicuculline and pentylenetetrazol. There were no significant effects on any type of strychnine seizure. Injections in the surrounding brainstem structures, including the pedunculopontine tegmental nucleus, had little effect on spontaneous behavior and did not significantly alter the thresholds of pentylenetetrazol-induced seizures. We have previously demonstrated that injections of GABA agonists in the central medial intralaminar nucleus of the thalamus have similar effects on behavior and seizures. Since the central medial nucleus receives important direct cholinergic projections from the laterodorsal tegmental nucleus, these two nuclei form a discrete ascending system which regulates seizure threshold.     

The dopamine-gamma aminobutyric acid interaction in the striatum of the rat is differently regulated by dopamine D-1 and D-2 types of receptor: evidence obtained with rotational behavioural experiments

The effects of GABA antagonists on apomorphine- and pergolide-induced rotational behaviour were studied with models combining intracerebral and systemic pharmacological treatments. Whether given systemically or intrastriatally to 6-hydroxydopamine-lesioned rats, the GABA antagonist picrotoxin inhibited the rotational responses produced by s.c. administration of the dopamine (DA) D-1/D-2 agonist apomorphine while it enhanced the rotational behaviour produced by the DA D-2 agonist pergolide. Following unilateral injection of picrotoxin or bicuculline into the striatum of naive rats, apomorphine produced ispsilateral rotation, while pergolide produced contralateral rotation. These contrasting effects are compared to the behavioural responses produced by intracerebral administration of GABAergic drugs alone. Intrapallidal injection of picrotoxin produced contralateral rotational behaviour which was independent of pallido-nigral pathways. Contralateral rotation was also produced by GABA agonists, but only following intranigral injections. The results are discussed in terms of differences in the localization of DA D-1 and DA D-2 receptors on striatal GABAergic neurons. The DA D-2 receptor agonist pergolide may induce inhibition of striato-pallidal GABAergic neurons, as well as of a local GABAergic circuit exerting inhibition on a striato--pallidal enkephalinergic pathway. However, the DA D-1/D-2 receptor agonist apomorphine may inhibit striatal interneurons exerting inhibition on a striato-nigral GABAergic projection. Such a neuronal arrangement may explain that striatal DA stimulation increases GABA release from the striato-nigral terminals.     

The ascending reticular activating system--from aminergic neurons to nitric oxide

The cholinergic neurons of the laterodorsal and pedunculopontine tegmental neurons are thought to comprise an important portion of the ascending reticular activating system. More recent work has demonstrated that the neurons of this cell group also released a number of neruoactive peptides and can produce nitric oxide in response to increases in intracellular calcium. The release of NO from the nerve terminals of these cells within the thalamus varies with behavioural state, being much lower during slow wave sleep than during wake and paradoxical sleep states. The NO release in the thalamus appears to act via the type II cGMP-dependent protein kinase present at high levels in the thalamic neurons. Thus the NO-cGMP signal transduction system can play an important role in regulating thalamic activity across behavioural states.     

Thalamic nucleus submedius receives GABAergic projection from thalamic reticular nucleus in the rat

GABAergic projection from thalamic reticular nucleus to thalamic nucleus submedius in the medial thalamus of the rat was studied by using immunohistochemistry for GABA, retrograde labeling with Fluoro-Gold combined with immunohistochemistry for GABA, and anterograde labeling with biotinylated dextranamine. Immunohistochemistry displayed that only GABA immunoreactive terminals were observed in the thalamic nucleus submedius, while GABA immunoreactive neuronal cell bodies were located in the thalamic reticular nucleus and lateral geniculate nucleus. Injection of Fluoro-Gold into the thalamic nucleus submedius resulted in massive retrogradely labeled neuronal cell bodies in the rostroventral portion of the ipsilateral thalamic reticular nucleus and a few in the contralateral thalamic reticular nucleus, and most of these cell bodies showed GABA immunopositive staining. Many biotinylated dextranamine anterogradely labeled fibers and terminals in the thalamic nucleus submedius were observed after injection of biotinylated dextranamine into the thalamic reticular nucleus. The present results provide a morphological evidence for a hypothesis that a disinhibitory effect on output neurons elicited by opioid or 5-hydroxytryptamine inhibiting a GABAergic terminal in the thalamic nucleus submedius may lead to activation of the descending inhibitory system and depression of the nociceptive inputs at the spinal cord level.     

Nicotine withdrawal hyperalgesia and opioid-mediated analgesia depend on nicotine receptors in nucleus accumbens

This study evaluated the capacity of mu-opioid and glutamate receptor agonists to differentially regulate the involvement of the GABAergic projection from the ventral pallidum to the mediodorsal thalamus in working memory and locomotor activity. Microinjection of either the ionotropic glutamate receptor agonist alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) or the mu agonist [D-Ala(2),N-Me-Phe(4),Gly-ol(5)]enkephalin into the ventral pallidum of male Sprague-Dawley rats produced a dose-dependent impairment in working memory, estimated using a forced delayed alternation task in a T-maze. Performance in a spatial discrimination task without delay was also impaired by glutamate, but not by mu receptor, stimulation. Involvement of the GABAergic projection from the ventral pallidum to the mediodorsal thalamus in mu-opioid-induced impairment of working memory was verified by showing that inhibiting GABA(B) receptors in the mediodorsal thalamus blocked the effect of [D-Ala(2),N-Me-Phe(4),Gly-ol(5)]enkephalin in the ventral pallidum. Similarly, either glutamate or mu-opioid receptor stimulation in the ventral pallidum elicited motor activity, and the motor stimulant effect of the mu agonist was blocked, while that of AMPA is not affected by GABA(B) receptor blockade in the mediodorsal thalamus. Distinction between mu and glutamate receptor stimulation was further revealed by the fact that stimulating mu receptors in the ventral pallidum caused a dose-dependent reduction in extracellular GABA levels, while AMPA was without effect on GABA in the ventral pallidum.These data indicate that stimulating mu-opioid receptors reduces GABAergic tone in the ventral pallidum, which increases activity in the GABAergic projection to the mediodorsal thalamus, thereby impairing working memory. Moreover, it is hypothesized that mu receptors in the ventral pallidum gate the recruitment of working memory into ongoing behavioral activity.     

Muscarinic receptors mediate enhancement of spontaneous GABA release in the chick brain

The functional role of muscarinic acetylcholine receptors in the lateral spiriform nucleus was studied in chick brain slices. Whole-cell patch-clamp recordings of neurons in the lateral spiriform nucleus revealed that carbachol enhanced GABAergic spontaneous inhibitory postsynaptic currents. The duration of the response to carbachol was significantly reduced after blockade of muscarinic receptors with atropine. In the presence of the nicotinic receptor antagonist dihydro-β-erythroidine, carbachol produced a delayed but prolonged enhancement of spontaneous GABAergic inhibitory postsynaptic currents that was completely blocked by atropine. Muscarine also enhanced the frequency of spontaneous GABAergic inhibitory postsynaptic currents in a dose-dependent manner, but had no effect on inhibitory postsynaptic current amplitude. While 4-diphenylacetoxy-N-(2-chloroethyl)-piperidine hydrochloride, a M₃ antagonist, completely blocked muscarine's effect, telenzepine, a M₁ antagonist, and tropicamide, a M₄ antagonist, only partially decreased the response to muscarine. Pirenzepine, a M₁ antagonist, and methoctramine, a M₂ antagonist, potentiated muscarine's enhancement of spontaneous GABAergic inhibitory postsynaptic currents. Muscarine's action was blocked by tetrodotoxin, cadmium chloride and ω-conotoxin GVIA, but was not affected by dihydro-β-erythroidine, 6-cyano-7-nitroquinoxaline-2,3-dione, d(−)-2-amino-5-phosphonopentanoic acid, naloxone or fluphenazine.These results demonstrate that activation of both muscarinic and nicotinic acetylcholine receptors can enhance GABAergic inhibitory postsynaptic currents in the lateral spiriform nucleus. The muscarinic response has a slower onset but lasts longer than the nicotinic effect. The M₃ receptor subtype is predominantly involved in enhancing spontaneous GABAergic inhibitory postsynaptic currents. These M₃ receptors must be located some distance from GABA release sites, since activation of voltage-dependent sodium channels, and consequent activation of N-type voltage-dependent calcium channels, is required to trigger enhanced GABA release following activation of muscarinic receptors.     

Dopamine inhibits GABA transmission from the globus pallidus to the thalamic reticular nucleus via presynaptic D4 receptors

The globus pallidus sends a significant GABAergic projection to the thalamic reticular nucleus. Because pallidal neurons express D4-dopamine receptors, we have explored their presence on pallidoreticular terminals by studying the effect of dopamine and D4-receptor agonists on the GABAergic transmission in the thalamic reticular nucleus. We made whole-cell recordings of inhibitory postsynaptic currents (IPSCs) and miniature inhibitory postsynaptic currents (mIPSCs) in the thalamic reticular neurons. Dopamine consistently reduced the IPSCs. The effect of dopamine was associated with paired-pulse facilitation, indicating a presynaptic location of the receptors. The effect of dopamine was also measured on the mIPSCs, reducing their frequency but not affecting their amplitude, which also suggests a presynaptic site of action. The selective D4-receptor agonist PD 168,077 also reduced the IPSCs, which was also associated with paired-pulse facilitation. In addition, this agonist reduced the frequency of the mIPSCs with no effect on their amplitude. The D4-receptor antagonist l-745,870 totally blocked the effect of the D4-receptor agonist, indicating the specificity of its effect. To verify the location of the receptors on the pallidal terminals, these were eliminated by injecting kainic acid into the globus pallidus. Kainic acid produced a drastic (80%) fall in the globus pallidus neuronal population. In this condition, the effect of the activation of D4 receptors both on the IPSCs and mIPSCs was prevented, thus indicating that the location of the receptors was on the pallidal terminals. Our results demonstrate that dopamine controls the activity of the thalamic reticular neurons by regulating the inhibitory input from the globus pallidus.     

5-HT1B receptor-mediated presynaptic inhibition of GABA release in the suprachiasmatic nucleus

The suprachiasmatic nucleus (SCN) receives a dense serotonergic innervation that modulates photic input to the SCN via serotonin 1B (5-HT1B) presynaptic receptors on retinal glutamatergic terminals. However, the majority of 5-HT1B binding sites in the SCN are located on nonretinal terminals and most axonal terminals in the SCN are GABAergic. We therefore tested the hypothesis that 5-HT1B receptors might also be located on SCN GABAergic terminals by examining the effects of the highly selective 5-HT1B receptor agonist CP-93,129 on SCN miniature inhibitory postsynaptic currents (mIPSCs). Whole cell patch-clamp recordings of mIPSCs were obtained from rat and mouse SCN neurons in hypothalamic slices. Using CsCl-containing microelectrodes with QX314, we isolated mPSCs that were sensitive to the GABAA receptor antagonist, bicuculline. Bath application of CP-93,129 (1 microM) decreased the frequency of mIPSCs by an average of 22% (n = 7) in rat SCN neurons and by an average of 30% (n = 8) in mouse SCN neurons with no clear effect on mIPSC amplitude. In mice lacking functional 5-HT1B receptors, CP-93,129 (1 microM) had no clear effect on the frequency or the amplitude of mIPSCs recorded in any of the cells tested (n = 4). The decrease in the frequency of mIPSCs of SCN neurons produced by the selective 5-HT1B receptor agonist CP-93,129 is consistent with the interpretation that 5-HT1B receptors are located on GABA terminals in the SCN and that 5-HT inhibits GABA release via a 5-HT1B presynaptic receptor-mediated mechanism.     

Adenosine A1 receptors control dopamine D1-dependent [(3)H]GABA release in slices of substantia nigra pars reticulata and motor behavior in the rat

Abnormalities in dopaminergic control of basal ganglia function play a key role in Parkinson's disease. Adenosine appears to modulate the dopaminergic control in striatum, where an inhibitory interaction between adenosine and dopamine receptors has been demonstrated. However the interaction has not been established in substantia nigra pars reticulata (SNr) where density of both receptors is high. Here we have explored the interaction between A1/D1 receptors in SNr. In SNr slices, SKF 38393, a selective D1 receptor agonist, produced a stimulation of depolarization-induced Ca(2+)-dependent [(3)H]GABA release that was inhibited by adenosine. The adenosine inhibition was abolished by 8-cyclopentyl-1,3-dipropylxanthine (DPCPX), a selective adenosine A1 receptor antagonist. DPCPX per se enhanced GABA release, indicating inhibition of the release by endogenous adenosine. When D1 receptors were blocked with SCH 23390 or the slices were depleted of dopamine, the effect of DPCPX was suppressed, showing that activation of dopamine receptors was necessary for the adenosine inhibition. In normal slices, 2-chloro-n(6)-cyclopentyladenosine (CCPA), a selective A1 agonist, inhibited GABA release, but the inhibition was prevented by the blockade of D1 receptors with SCH 23390. Superperfusion with 8-bromo-cAMP produced a stimulation of GABA release that was not blocked by CCPA: this finding indicates that the blockade of D1 effects caused by activation of A1 receptors is specific. To see if these actions on GABA release were correlated with changes in motor behavior we studied the effect of unilateral intranigral injections of modifiers of adenosine A1 and dopamine D1 receptors in rats challenged with systemic methamphetamine. Both the A1 agonist CCPA and the D1 antagonist SCH 23390 produced ipsilateral turning whereas the A1 antagonist DPCPX caused contralateral turning. These motor effects are consistent with the findings on GABA release.The results indicate the presence of an inhibitory A1/D1 receptor interaction in SNr. The inhibition exerted by A1 adenosine receptors on GABAergic striatonigral transmission would be due exclusively to blockade of the facilitation resulting from activation of D1 dopamine receptors. The data permit to better understand the action of adenosine antagonists in the treatment of Parkinson's disease.