Spontaneous GABAergic Inhibitory Postsynaptic Currents Research Articles

Nicotine increases GABAergic input on rat dorsal raphe serotonergic neurons through alpha7 nicotinic acetylcholine receptor.

The dorsal raphe nucleus (DRN) contains large populations of serotonergic (5-HT) neurons. This nucleus receives GABAergic inhibitory afferents from many brain areas and from DRN interneurons. Both GABAergic and 5-HT DRN neurons express functional nicotinic acetylcholine receptors (nAChRs). Previous studies have demonstrated that nicotine increases 5-HT release and 5-HT DRN neuron discharge rate by stimulating postsynaptic nAChRs and by increasing glutamate and norepinephrine release inside DRN. However, the influence of nicotine on the GABAergic input to 5-HT DRN neurons was poorly investigated. Therefore, the aim of this work was to determine the effect of nicotine on GABAergic spontaneous inhibitory postsynaptic currents (sIPSCs) of 5-HT DRN neurons and the subtype of nAChR(s) involved in this response. Experiments were performed in coronal slices obtained from young Wistar rats. GABAergic sIPSCs were recorded from post hoc-identified 5-HT DRN neurons with the whole cell voltage patch-clamp technique. Administration of nicotine (1 μM) increased sIPSC frequency in 72% of identified 5-HT DRN neurons. This effect was not reproduced by the α4β2 nAChR agonist RJR-2403 and was not influenced by TTX (1 μM). It was mimicked by the selective agonist for α7 nAChR, PNU-282987, and exacerbated by the positive allosteric modulator of the same receptor, PNU-120596. The nicotine-induced increase in sIPSC frequency was independent on voltage-gated calcium channels and dependent on Ca(2+)-induced Ca(2+) release (CICR). These results demonstrate that nicotine increases the GABAergic input to most 5-HT DRN neurons, by activating α7 nAChRs and producing CICR in DRN GABAergic terminals.

Serotonergic modulation of inhibitory synaptic transmission in mouse inferiorcolliculus

The inferior colliculus (IC) transmits the ascending auditory signal to the thalamic medial geniculate nucleus. Previous studies have reported that serotonergic input originating from the raphe nuclei has a strong influence on signal processing within the central nucleus of the IC. To identify the cellular target for the serotonergic modulation in the IC, we examined the effect of serotonin as well as selective serotonin reuptake inhibitor (SSRI) fluvoxamine on spontaneous GABAergic and glycinergic inhibitory postsynaptic currents (sIPSCs) recorded with whole-cell recordings.Consistent with earlier studies, we confirmed that serotonin robustly enhanced the frequency, but not amplitude, of GABAergic sIPSCs. It should be noted that the application of fluvoxamine alone marginally increased the frequency of GABAergic sIPSCs. These findings suggest that serotonin is endogenously released even in slice preparations, and it negatively modulates the tone of activity of inhibitory neurons within IC. We also examined the effect of serotonin and fluvoxamine on glycinergic sIPSCs and found that serotonin has a significantly weaker effect on glycinergic sIPSCs than on GABAergic sIPSCs. The differential sensitivity of the GABAergic and glycinergic sIPSCs to serotonin implies that serotonergic input plays a specific role in auditory information processing.Moreover, it suggests that the serotonergic input may contribute to pathological conditions such as tinnitus.

Propofol Stimulates Noradrenalin-Inhibited Neurons in the Ventrolateral Preoptic Nucleus by Reducing GABAergic Inhibition

The cellular mechanisms underlying the sedative effect of general anesthetics are not completely understood. Accumulating evidence indicates that the ventrolateral preoptic area (VLPO) of the hypothalamus plays a critical role. The VLPO contains 2 major types of neurons, the noradrenalin-inhibited GABAergic projecting neurons (NA(-) neurons) and the noradrenalin-excited interneurons (NA(+) neurons) which are probably also γ-aminobutyric acid (GABA)-containing neurons. Our previous work suggests that NA(-) neurons are normally under the inhibitory control of NA(+) neurons. Previous studies also show that GABAergic agents including propofol activate GABAergic projecting neurons in the VLPO, which is believed to lead to the inhibition of the arousal-producing nuclei in the tuberomammillary nucleus and sedation. However, how propofol activates VLPO neurons remains unclear. We explored the possibility that propofol activates NA(-) neurons indirectly, by inhibiting GABAergic transmission including those from VLPO NA(+) neurons. Electrophysiological activities were recorded from VLPO cells in acute brain slices of rats. Propofol facilitates the discharges of NA(-) neurons and reduces the frequency, but not the amplitude of spontaneous GABAergic inhibitory postsynaptic currents in NA(-) neurons. Conversely, propofol suppressed the discharges of NA(+) neurons. Propofol excites VLPO NA(-) neurons by reducing GABAergic transmission, at least in part by inhibiting VLPO NA(+) neurons. This may be a critical mechanism contributing to propofol-induced sedation.

The effects of behavioral control over stress on GABAergic spontaneous inhibitory postsynaptic currents in prefrontal cortical pyramidal neurons

Traumatic events may lead to anxiety, depression and post-traumatic stress disorder (PTSD). However, the majority of individuals exposed to trauma do not develop these disorders. The stressor controllability paradigm has been widely used as a model for understanding the neurobiology underlying factors that confer vulnerability and resilience to the outcome of traumatic events. In this paradigm rats receive a series of tail shocks: one group of rats have control over the termination of the shock by means of turning a wheel (escapable shock, ES), while the other “yoked” group of rats receive physically identical shocks but have no control over shock termination (inescapable shock, IS). In subsequent behavioral tests that model components of anxiety and depression, IS rats without control show increased signs of behavioral depression, while ES rats that have control over the shock behave as naïve home caged (HC) rats. We have previously reported that individual deep layer pyramidal neurons from the ventral medial prefrontal cortex (vmPFC) exhibit changes in their intrinsic excitability following ES. To examine if there is a corresponding reduction in synaptic inhibition, we tested IS, ES and HC deep layer pyramidal neurons under identical conditions. Collecting such electrophysiological data from pyramidal neurons after exposure to stress is a technical challenge, yet very useful for conductance-based neural simulations and computational modeling. Here we present a data set of spontaneous inhibitory postsynaptic currents (sIPSCs) gathered from whole-cell patch-clamp recordings of individual prefrontal cortical deep layer neurons from adult rats (60-70 days old) after exposure to ES, IS or HC. In order to analyze the data, we provide our script used for the detection of synaptic events written for the scientific/engineering program Igor Pro that allows users to define their own event detection parameters.

Developmental Regulation and Activity-Dependent Maintenance of GABAergic Presynaptic Inhibition onto Rod Bipolar Cell Axonal Terminals

Presynaptic inhibition onto axons regulates neuronal output, but how such inhibitory synapses develop and are maintained invivo remains unclear. Axon terminals of glutamatergic retinal rod bipolar cells (RBCs) receive GABAA and GABAC receptor-mediated synaptic inhibition. We found that perturbing GABAergic or glutamatergic neurotransmission does not prevent GABAergic synaptogenesis onto RBC axons. But, GABA release is necessary for maintaining axonal GABA receptors. This activity-dependent process is receptor subtype specific: GABAC receptors are maintained, whereas GABAA receptors containing α1, but not α3, subunits decrease over time in mice with deficient GABA synthesis. GABAA receptor distribution on RBC axons is unaffected in GABAC receptor knockout mice. Thus, GABAA and GABAC receptor maintenance are regulated separately. Although immature RBCs elevate their glutamate release when GABA synthesis is impaired, homeostatic mechanisms ensure that the RBC output operates within its normal range after eye opening, perhaps to regain proper visual processing within the scotopic pathway.

Inflammation Subverts Hippocampal Synaptic Plasticity in Experimental Multiple Sclerosis

Abnormal use-dependent synaptic plasticity is universally accepted as the main physiological correlate of memory deficits in neurodegenerative disorders. It is unclear whether synaptic plasticity deficits take place during neuroinflammatory diseases, such as multiple sclerosis (MS) and its mouse model, experimental autoimmune encephalomyelitis (EAE). In EAE mice, we found significant alterations of synaptic plasticity rules in the hippocampus. When compared to control mice, in fact, hippocampal long-term potentiation (LTP) induction was favored over long-term depression (LTD) in EAE, as shown by a significant rightward shift in the frequency–synaptic response function. Notably, LTP induction was also enhanced in hippocampal slices from control mice following interleukin-1β (IL-1β) perfusion, and both EAE and IL-1β inhibited GABAergic spontaneous inhibitory postsynaptic currents (sIPSC) without affecting glutamatergic transmission and AMPA/NMDA ratio. EAE was also associated with selective loss of GABAergic interneurons and with reduced gamma-frequency oscillations in the CA1 region of the hippocampus. Finally, we provided evidence that microglial activation in the EAE hippocampus was associated with IL-1β expression, and hippocampal slices from control mice incubated with activated microglia displayed alterations of GABAergic transmission similar to those seen in EAE brains, through a mechanism dependent on enhanced IL-1β signaling. These data may yield novel insights into the basis of cognitive deficits in EAE and possibly of MS.

Social Stress Alters Inhibitory Synaptic Input to Distinct Subpopulations of Raphe Serotonin Neurons

Anxiety disorders are among the most prevalent psychiatric disorders, yet much is unknown about the underlying mechanisms. The dorsal raphe (DR) is at the crux of the anxiety-inducing effects of uncontrollable stress, a key component of models of anxiety. Though DR serotonin (5-HT) neurons play a prominent role, anxiety-associated changes in the physiology of 5-HT neurons remain poorly understood. A 5-day social defeat model of anxiety produced a multifaceted, anxious phenotype in intruder mice that included increased avoidance behavior in the open field test, increased stress-evoked grooming, and increased bladder and heart weights when compared to control mice. Intruders were further compared to controls using electrophysiology recordings conducted in midbrain slices wherein recordings targeted 5-HT neurons of the ventromedial (vmDR) and lateral wing (lwDR) subfields of the DR. Though defining membrane characteristics of 5-HT neurons were unchanged, γ-aminobutyric-acid-mediated (GABAergic) synaptic regulation of 5-HT neurons was altered in a topographically specific way. In the vmDR of intruders, there was a decrease in the frequency and amplitude of GABAergic spontaneous inhibitory postsynaptic currents (sIPSCs). However, in the lwDR, there was an increase in the strength of inhibitory signals due to slower sIPSC kinetics. Synaptic changes were selective for GABAergic input, as glutamatergic synaptic input was unchanged in intruders. The distinct inhibitory regulation of DR subfields provides a mechanism for increased 5-HT output in vmDR target regions and decreased 5-HT output in lwDR target regions, divergent responses to uncontrollable stress that have been reported in the literature but were previously poorly understood.

Muscarinic M4 receptors regulate GABAergic transmission in rat tuberomammillary nucleus neurons

Histaminergic neurons within the tuberomammillary nucleus (TMN) play an important role in sleep-wakefulness regulation. Here, we report the muscarinic modulation of GABAergic spontaneous miniature inhibitory postsynaptic currents (mIPSCs) in mechanically dissociated rat histaminergic neurons using a conventional whole-cell patch clamp technique. Muscarine, a nonselective muscarinic acetylcholine (mACh) receptor agonist, reversibly decreased mIPSC frequency without affecting the current amplitude, indicating that muscarine acts presynaptically to decrease the probability of spontaneous GABA release. The muscarine action on GABAergic mIPSC frequency was completely blocked by atropine, a nonselective mACh receptor antagonist, and tropicamide, an M4 receptor antagonist. The muscarine-induced decrease in mIPSC frequency was completely occluded in the presence of Cd2+, a general voltage-dependent Ca2+ channel blocker, or in a Ca2+-free external solution. However, pharmacological agents affecting adenylyl cyclase or G-protein coupled inwardly rectifying K+ channel activity did not prevent the inhibitory action of muscarine on GABAergic mIPSCs. These results suggest that muscarine acts on M4 receptors on GABAergic nerve terminals projecting to histaminergic neurons to inhibit spontaneous GABA release via the inhibition of Ca2+ influx from the extracellular space. Muscarine also inhibited action potential-dependent GABA release by activating presynaptic M4 receptors in more physiological conditions. The M4 receptor-mediated modulation of GABAergic transmission onto TMN neurons may contribute to the regulation of sleep-wakefulness.

GABAergic Actions Mediate Opposite Ethanol Effects on Dopaminergic Neurons in the Anterior and Posterior Ventral Tegmental Area

It is known that the posterior ventral tegmental area (p-VTA) differs from the anterior VTA (a-VTA) in that rats learn to self-administer ethanol into the p-VTA, but not into the a-VTA. Because activation of VTA dopaminergic neurons by ethanol is a cellular mechanism underlying the reinforcement of ethanol consumption, we hypothesized that ethanol may exert different effects on dopaminergic neurons in the p-VTA and a-VTA. In patch-clamp recordings in midbrain slices from young rats (postnatal days 22-32), we detected no significant difference in electrophysiological properties between p-VTA and a-VTA dopaminergic neurons. However, acute exposure to ethanol (21-86 mM) stimulated p-VTA dopaminergic neurons but suppressed a-VTA dopaminergic neurons. Conversely, ethanol (>21 mM) dose-dependently reduced the frequency of the GABAergic spontaneous inhibitory postsynaptic currents (sIPSCs) generated by inhibitory neuronal firing but not miniature inhibitory postsynaptic currents (mIPSCs) in p-VTA dopaminergic neurons. By contrast, ethanol increased the frequency and amplitude of both sIPSCs and mIPSCs in a-VTA dopaminergic neurons. All of these effects of ethanol were abolished by a GABA(A) receptor antagonist. There was a strong negative correlation between ethanol-evoked modulation of sIPSCs and neuronal firing in VTA dopaminergic neurons. These results indicate that GABAergic inputs play an important role in ethanol's actions in the VTA. The differential effects of ethanol on sIPSCs and neuronal firing in the p-VTA and a-VTA could be the basis for ethanol reinforcement via the p-VTA.

Exogenous and Endogenous Cannabinoids Suppress Inhibitory Neurotransmission in the Human Neocortex

Activation of CB(1) receptors on axon terminals by exogenous cannabinoids (eg, Δ(9)-tetrahydrocannabinol) and by endogenous cannabinoids (endocannabinoids) released by postsynaptic neurons leads to presynaptic inhibition of neurotransmission. The aim of this study was to characterize the effect of cannabinoids on GABAergic synaptic transmission in the human neocortex. Brain slices were prepared from neocortical tissues surgically removed to eliminate epileptogenic foci. Spontaneous GABAergic inhibitory postsynaptic currents (sIPSCs) were recorded in putative pyramidal neurons using patch-clamp techniques. To enhance the activity of cannabinoid-sensitive presynaptic axons, muscarinic receptors were continuously stimulated by carbachol. The synthetic cannabinoid receptor agonist WIN55212-2 decreased the cumulative amplitude of sIPSCs. The CB(1) antagonist rimonabant prevented this effect, verifying the involvement of CB(1) receptors. WIN55212-2 decreased the frequency of miniature IPSCs (mIPSCs) recorded in the presence of tetrodotoxin, but did not change their amplitude, indicating that the neurotransmission was inhibited presynaptically. Depolarization of postsynaptic pyramidal neurons induced a suppression of sIPSCs. As rimonabant prevented this suppression, it is very likely that it was due to endocannabinods acting on CB(1) receptors. This is the first demonstration that an exogenous cannabinoid inhibits synaptic transmission in the human neocortex and that endocannabinoids released by postsynaptic neurons suppress synaptic transmission in the human brain. Interferences of cannabinoid agonists and antagonists with synaptic transmission in the cortex may explain the cognitive and memory deficits elicited by these drugs.

Endogenous BDNF regulates inhibitory synaptic transmission in the ventromedial nucleus of the hypothalamus

Output from steroidogenic factor-1 (SF-1) neurons in the ventromedial nucleus of the hypothalamus (VMH) is anorexigenic. SF-1 neurons express brain-derived neurotrophic factor (BDNF) that contributes to the regulation of food intake and body weight. Here I show that regulation of GABAergic inputs onto SF-1 neurons by endogenous BDNF determines the anorexigenic outcome from the VMH. Single-cell RT-PCR analysis reveals that one-third of SF-1 neurons express BDNF and that only a subset of BDNF-expressing SF-1 neurons coexpresses the melanocortin receptor type 4. Whole cell patch-clamp analysis of SF-1 neurons in the VMH shows that exogenous BDNF significantly increases the frequency of spontaneous GABAergic inhibitory postsynaptic currents (sIPSCs). This enhancement of GABA drive readily decreases the excitability of SF-1 neurons. However, treatment with BDNF has no significant effect on the frequency of TTX-independent GABAergic IPSCs. Moreover, TrkB receptors are not localized at the postsynaptic sites of GABAergic synapses on SF-1 neurons as there is no change in the amplitude of miniature IPSCs in the presence of BDNF. Dual patch-clamp recordings in mouse hypothalamic slices reveal that stimulation of one SF-1 neuron induces an increase in sIPSC frequency onto the neighboring SF-1 neuron. More importantly, this effect is blocked by a tyrosine kinase inhibitor. Hence, this increased GABA drive onto SF-1 neurons may, in part, explain the cellular mechanisms that mediate the anorexigenic effects of BDNF.

A neuroprotective role for polyamines in a Xenopus tadpole model of epilepsy

Polyamines are endogenous molecules involved in cell damage following neurological insults, although it is unclear whether polyamines reduce or exacerbate this damage. We used a developmental seizure model in which we exposed Xenopus laevis tadpoles to pentylenetetrazole (PTZ), a known convulsant. We found that, after an initial PTZ exposure, seizure onset times were delayed in response to a second PTZ exposure 4 h later. This protective effect was a result of activity-dependent increases in synthesis of putrescine, the simplest polyamine. Unlike more complex polyamines that directly modulate ion channels, putrescine exerted its effect by altering the balance of excitation to inhibition. Tectal neuron recordings, 4 h after the initial seizure, revealed an elevated frequency of GABAergic spontaneous inhibitory postsynaptic currents. Our data suggest that this effect is mediated by an atypical pathway that converts putrescine into GABA, which then activates presynaptic GABA(B) receptors. Our data suggest that polyamines have a previously unknown neuroprotective role in the developing brain.

Multiple effects of allopregnanolone on GABAergic responses in single hippocampal CA3 pyramidal neurons

3α-Hydroxy, 5α-reduced pregnane steroids, such as allopregnanolone, are potent modulators of GABA A receptors and have many biological responses including sedative, anxiolytic, anticonvulsant and anesthetic actions. In the present study, we have investigated the effects of allopregnanolone on GABA A receptors in acutely isolated single hippocampal CA3 pyramidal neurons using the whole cell patch-clamp technique. Allopregnanolone induced membrane Cl − currents in a concentration-dependent manner, and the allopregnanolone-induced currents (I AlloP) were blocked by noncompetitive GABA A receptor antagonists. The I AlloP was not affected by the intracellular loading of γ-cyclodextrin (γ-CD), which efficiently sequesters several kinds of endogenous neurosteroids including allopregnanolone, suggesting that allopregnanolone accesses extracellular but not intracellular sites to activate GABA A receptors. Allopregnanolone prolonged the decay time constant of GABAergic spontaneous inhibitory postsynaptic currents (sIPSCs), suggesting that allopregnanolone modulates the desensitization kinetics of postsynaptic GABA A receptors. The picrotoxin-sensitive tonic currents (I tonic), which were mediated by extrasynaptic GABA A receptors, were recorded from CA3 pyramidal neurons. The intracellular loading of γ-CD or allopregnanolone significantly decreased or increased the amplitude of picrotoxin-sensitive I tonic, respectively, suggesting that endogenous neurosteroids might, at least in part, be involved in the generation of picrotoxin-sensitive I tonic. Allopregnanolone also increased the frequency of GABAergic sIPSCs, in a manner dependent on the integrity of voltage-dependent Na + and Ca 2+ channels, suggesting that allopregnanolone activates presynaptic GABA A receptors to depolarize GABAergic nerve terminals. The present results suggest that allopregnanolone exerts its pharmacological and pathophysiological actions via the modulation of multiple types of GABA A receptor-mediated responses.

A New Kv1.2 Channelopathy Underlying Cerebellar Ataxia

A forward genetic screen of mice treated with the mutagen ENU identified a mutant mouse with chronic motor incoordination. This mutant, named Pingu (Pgu), carries a missense mutation, an I402T substitution in the S6 segment of the voltage-gated potassium channel Kcna2. The gene Kcna2 encodes the voltage-gated potassium channel α-subunit Kv1.2, which is abundantly expressed in the large axon terminals of basket cells that make powerful axo-somatic synapses onto Purkinje cells. Patch clamp recordings from cerebellar slices revealed an increased frequency and amplitude of spontaneous GABAergic inhibitory postsynaptic currents and reduced action potential firing frequency in Purkinje cells, suggesting that an increase in GABA release from basket cells is involved in the motor incoordination in Pgu mice. In line with immunochemical analyses showing a significant reduction in the expression of Kv1 channels in the basket cell terminals of Pgu mice, expression of homomeric and heteromeric channels containing the Kv1.2(I402T) α-subunit in cultured CHO cells revealed subtle changes in biophysical properties but a dramatic decrease in the amount of functional Kv1 channels. Pharmacological treatment with acetazolamide or transgenic complementation with wild-type Kcna2 cDNA partially rescued the motor incoordination in Pgu mice. These results suggest that independent of known mutations in Kcna1 encoding Kv1.1, Kcna2 mutations may be important molecular correlates underlying human cerebellar ataxic disease.

Loss of Leptin Receptors on Hypothalamic POMC Neurons Alters Synaptic Inhibition

Adaptive changes in hypothalamic neural circuitry occur in response to alterations in nutritional status. This plasticity at hypothalamic synapses contributes to the control of food intake and body weight. Here we show that genetic ablation of leptin receptor gene expression in pro-opiomelanocortin (POMC) neurons (POMC: Lepr(-/-) GFP) induces alterations at synapses on POMC neurons in the arcuate nucleus of the hypothalamus. Our studies reveal that POMC: Lepr(-/-) GFP mice have decreased frequency of spontaneous GABAergic, but not glutamatergic, postsynaptic currents at synapses on POMC neurons. The decay time course of GABAergic spontaneous inhibitory postsynaptic currents (sIPSCs) onto POMC neurons in POMC: Lepr(-/-) GFP mice is significantly slower than that of sIPSCs in control animals. While analysis of individual miniature IPSCs shows lowered baseline activity, this tonic decrease is associated with an increased amplitude and slow decay of mini-IPSCs onto POMC neurons in POMC: Lepr(-/-) GFP mice. Moreover, POMC neurons receive greater total ionic flux per GABAergic event in the absence of leptin receptor signaling. In addition, treatment with the alpha 3 subunit-containing GABA(A) receptor modulator SB-205384 enhances GABAergic transmission only onto POMC neurons in POMC: Lepr(-/-) GFP mice. Single-cell RT-PCR analysis further supports the expression of the alpha 3 subunit of the GABA(A) receptor on POMC neurons in POMC: Lepr(-/-) GFP mice. Finally, the responses to the GABA(A) receptor agonist isoguvacine of POMC neurons are significantly smaller in POMC: Lepr(-/-) GFP than in control animals. Therefore our present work demonstrates that loss of leptin signaling in POMC neurons induces synaptic alterations at POMC synapses that may play an essential role in energy homeostasis.

Activation of serotonin 1A receptors in ventrolateral orbital cortex depresses persistent nociception: A presynaptic inhibition mechanism

The present study examined the effect of serotonin 1A (5-HT 1A) receptor activation in the ventrolateral orbital cortex (VLO) upon formalin-evoked flinching behavior and spinal Fos expression, and further determined whether activation of 5-HT 1A receptors affected the spontaneous GABAergic miniature inhibitory postsynaptic currents (mIPSCs) in rat VLO slice by pharmacologically separated neurons to understand the possible mechanism underlying this effect. Microinjection of the 5-HT 1A receptors agonist 8-OH-DPAT (8-hydro-2-(di-n-propylamino) tetralin) into the VLO depressed the formalin-evoked nociceptive behavior flinching response and the Fos expression in the lumbar spinal cord dorsal, which was antagonized by pre-treatment with 5-HT 1A receptors antagonist NAN-190 (1-(2-methoxyphenyl)-4-[4-(2-phthalimido)butyl]piperazine hydrobromide). Furthermore, application of 8-OH-DPAT into VLO slice inhibited GABAergic mIPSC frequency in a dose-dependent manner without effects on amplitude of the GABAergic mIPSCs, this effect was blocked by NAN-190. These results provide evidence for the involvement of 5-HT 1A receptors in VLO in the modulation of persistent inflammatory nociception, and suggest that a presynaptic inhibition of the GABA release may contribute to the 5-HT 1A receptor-mediated descending antinociception.

Compound K, a metabolite of ginsenosides, facilitates spontaneous GABA release onto CA3 pyramidal neurons

Ginsenoside Rb1, a major ingredient of ginseng saponins, can affect various brain functions, including learning and memory. When ingested orally, ginsenoside Rb1 is not found in plasma as well as urine, but its metabolite compound K (ComK) reaches the systemic circulation in animals and human. Nevertheless, the pharmacological actions of ComK are still poorly known. In the present study, we investigated the effect of ComK on GABAergic spontaneous miniature inhibitory post-synaptic currents (mIPSCs) in acutely isolated rat hippocampal CA3 pyramidal neurons using a conventional whole-cell patch-clamp technique. While ComK significantly increased mIPSC frequency in a concentration-dependent manner, it had no effect on the current amplitude, suggesting that ComK acts pre-synaptically to increase the probability of spontaneous GABA release. ComK still increased mIPSC frequency even in a Ca(2+) -free external solution, suggesting that the ComK-induced increase spontaneous GABA release is not related to Ca(2+) influx from the extracellular space. However, the ComK-induced increase mIPSC frequency was significantly decreased after the blockade of either sarcoplasmic/endoplasmic reticulum Ca(2+) -ATPase or Ca(2+) release channels. These results strongly suggest that ComK enhances spontaneous GABA release by increasing intraterminal Ca(2+) concentration via Ca(2+) release from pre-synaptic Ca(2+) stores. The ComK-induced modulation of inhibitory transmission onto CA3 pyramidal neurons could have a broad impact on the excitability of CA3 pyramidal neurons and affect the physiological functions mediated by the hippocampus.

Presynaptic kainate receptors increase GABAergic neurotransmission in rat periaqueductal gray neurons

Neurons within the periaqueductal gray (PAG) have been implicated in the central regulation of pain signals by affecting the descending inhibitory pathway. Here we report on the functional role of presynaptic kainate receptors within the PAG. Using a conventional whole-cell patch clamp technique, we recorded GABAergic spontaneous miniature inhibitory postsynaptic currents (mIPSCs) from mechanically isolated rat PAG neurons in the presence of 300 nM tetrodotoxin and 20 µM dl-2-amino-5-phosphonovaleric acid under voltage-clamp conditions. Kainic acid at a 10 µM concentration significantly increased the frequency of GABAergic mIPSCs without affecting their amplitude, suggesting that kainic acid acts presynaptically to enhance spontaneous GABA release. The kainic acid-induced increase in mIPSC frequency was completely blocked by CNQX, a selective AMPA/kainate receptor antagonist. While neither AMPA nor NMDA affected GABAergic mIPSC frequency, ATPA, a selective agonist of GluR5-containing kainate receptors, increased GABAergic mIPSC frequency in a concentration-dependent manner. The kainic acid-induced increase in mIPSC frequency was completely suppressed either in the presence of 100 µM Cd 2+, a general voltage-dependent Ca 2+ channel (VDCC) blocker, or in the Na +-free external solution. These results suggest that presynaptic kainate receptors have a low permeability to Ca 2+, and that their activation elicits a presynaptic depolarization large enough to activate presynaptic VDCCs. Presynaptic kainate receptors on GABAergic nerve terminals appear to modulate GABAergic transmission, and in doing so may play an important role in the regulation of PAG neuron excitability.

Increase of GABA A receptor-mediated tonic inhibition in dentate granule cells after traumatic brain injury

Traumatic brain injury (TBI) can result in altered inhibitory neurotransmission, hippocampal dysfunction, and cognitive impairments. GABAergic spontaneous and miniature inhibitory postsynaptic currents (sIPSCs and mIPSCs) and tonic (extrasynaptic) whole cell currents were recorded in control rat hippocampal dentate granule cells (DGCs) and at 90 days after controlled cortical impact (CCI). At 34 °C, in CCI DGCs, sIPSC frequency and amplitude were unchanged, whereas mIPSC frequency was decreased (3.10 ± 0.84 Hz, n = 16, and 2.44 ± 0.67 Hz, n = 7, p < 0.05). At 23 °C, 300 nM diazepam increased peak amplitude of mIPSCs in control and CCI DGCs, but the increase was 20% higher in control (26.81 ± 2.2 pA and 42.60 ± 1.22 pA, n = 9, p = 0.031) compared to CCI DGCs (33.46 ± 2.98 pA and 46.13 ± 1.09 pA, n = 10, p = 0.047). At 34 °C, diazepam did not prolong decay time constants (6.59 ± 0.12 ms and 6.62 ± 0.98 ms, n = 9, p = 0.12), the latter suggesting that CCI resulted in benzodiazepine-insensitive pharmacology in synaptic GABA A receptors (GABA ARs). In CCI DGCs, peak amplitude of mIPSCs was inhibited by 100 μM furosemide (51.30 ± 0.80 pA at baseline and 43.50 ± 5.30 pA after furosemide, n = 5, p < 0.001), a noncompetitive antagonist of GABA ARs with an enhanced affinity to α4 subunit-containing receptors. Potentiation of tonic current by the GABA AR δ subunit-preferring competitive agonist THIP (1 and 3 µM) was increased in CCI DGCs (47% and 198%) compared to control DGCs (13% and 162%), suggesting the presence of larger tonic current in CCI DGCs; THIP (1 µM) had no effect on mIPSCs. Taken together, these results demonstrate alterations in synaptic and extrasynaptic GABA ARs in DGCs following CCI.

Tonic Modulation of GABA Release by Nicotinic Acetylcholine Receptors in Layer V of the Murine Prefrontal Cortex

By regulating the neocortical excitability, nicotinic acetylcholine receptors (nAChRs) control vigilance and cognition and are implicated in epileptogenesis. Modulation of gamma-aminobutyric acid (GABA) release often accompanies these processes. We studied how nAChRs regulate GABAergic transmission in the murine neocortex with immunocytochemical and patch-clamp methods. The cholinergic fibers densely innervated the somatosensory, visual, motor, and prefrontal cortices (PFC). Laminar distribution was broadly homogeneous, especially in the PFC. The cholinergic terminals were often adjacent to the soma and dendrites of GABAergic interneurons, but well-differentiated synapses were rare. Tonically applied nicotine (1-100 microM) increased the frequency of spontaneous GABAergic inhibitory postsynaptic currents (IPSCs) on pyramidal neurons in PFC layer V. The contribution of nAChR types was assessed by using 1 microM dihydro-beta-erythroidine (DHbetaE), to block heteromeric nAChRs, and 10 nM methyllycaconitine (MLA), to block homomeric nAChRs. Both inhibitors antagonized the effect of nicotine on IPSCs, suggesting that mixed nAChR types control pyramidal neuron inhibition in layer V. To determine whether nAChRs are expressed on basket cells' terminals, we studied miniature IPSCs (mIPSCs). These were revealed using 0.5 microM tetrodotoxin and 50 microM Cd(2+) to isolate the GABAergic terminals from the action potential drive. The nicotinic stimulation of mIPSCs was antagonized by DHbetaE, but not MLA, indicating that heteromeric nAChRs prevail in GABAergic terminals. Immunocytochemistry confirmed the expression of nAChRs on basket cells' somata and terminals. Finally, when the ionotropic glutamatergic transmission was blocked, nicotine partially inhibited the IPSCs, an effect counteracted by both DHbetaE and MLA. Therefore, a fraction of nAChRs are capable of activating GABAergic interneurons that in turn inhibit other GABAergic interneurons, thereby reducing the IPSCs. We conclude that heteromeric nAChRs control GABA release presynaptically, whereas mixed nAChRs regulate both excitation and inhibition of interneurons, the balance depending on the overall glutamatergic drive.