Sleep deprivation induces changes in 5-HT actions and 5-HT1A receptor expression in the rat hippocampus.

A 12h sleep deprivation enhances the expression of 5-hydroxytryptamine (5-HT) 1A receptors in rat hippocampus that recedes with 48h sleep recovery. The depressant effect of applied 5-HT on the field excitatory postsynaptic potentials recorded in the CA1 area, is also enhanced in hippocampi of SD rats. Following a 24 or 48h sleep recovery, the increase in the 5-HT effect subsided. These results have implications for therapeutics treating clinical depression.

Are presynaptic GABA-Cρ2 receptors involved in anti-nociception?

We investigated the anti-nociceptive effects of GABA-C receptors in the central nervous system. Intracisternal injection of CACA, a GABA-C receptor agonist or isoguvacine, a GABA-A receptor agonist, significantly increased the tail-withdrawal latency. TPMPA, a GABA-C receptor antagonist blocked the effects of CACA but not isoguvacine indicating that GABA-C receptors are involved in regulating pain. Further, double-labelled immunofluorescence studies revealed that GABA-Cρ2 receptors are expressed presynaptically in the spinal dorsal horn, especially, substantia gelatinosa, a region that has been previously implicated in analgesia by regulating nociceptive inflow. These data provide a provenance for future work looking at presynaptic spinal GABA-C receptors in the control of nociception.

Activity-mediated plasticity of GABA equilibrium potential in rat hippocampal CA1 neurons.

The equilibrium potential (E(GABA)(-PSC)) for gamma-aminobutyric acid (GABA) A receptor mediated inhibitory postsynaptic currents (PSCs) in hippocampal CA1 pyramidal neurons shifts when theta-burst stimulation (four pulses at 100 Hz in each burst in a train consisting of five bursts with an inter-burst interval of 200 ms, the train repeated thrice at 30-s intervals) is applied to the input. E(GABA)(-PSC) is regulated by K(+)/Cl(-) co-transporter (KCC2). GABA(B) receptors are implicated in modulating KCC2 levels. In the current study, the involvement of KCC2, as well as GABA(B) receptors, in theta-burst-mediated shifts in E(GABA)(-PSC) was examined. Whole-cell patch recordings were made from hippocampal CA1 pyramidal neurons (from 9 to 12 days old rats), in a slice preparation. Glutamatergic excitatory postsynaptic currents were blocked with dl-2-amino-5-phosphonovaleric acid (50 microM) and 6,7-dinitroquinoxaline-2,3-dione (20 microM). The PSC and the E(GABA)(-PSC) were stable when stimulated at 0.05 Hz. However, both changed following a 30-min stimulation at 0.5 or 1 Hz. Furosemide (500 microM) and KCC2 anti-sense in the recording pipette but not bumetanide (20 or 100 microM) or KCC2 sense, blocked the changes, suggesting KCC2 involvement. Theta-burst stimulation induced a negative shift in E(GABA)(-PSC), which was prevented by KCC2 anti-sense; however, KCC2 sense had no effect. CGP55845 (2 microM), a GABA(B) antagonist, applied in the superfusing medium, or GDP-beta-S in the recording pipette, blocked the shift in E(GABA)(-PSC). These results indicate that activity-mediated plasticity in E(GABA)(-PSC) occurs in hippocampal CA1 pyramidal neurons and theta-burst-induced negative shift in E(GABA)(-PSC) requires KCC2, GABA(B) receptors and G-protein activation.

Long-term depression of excitatory synaptic transmission in rat hippocampal CA1 neurons following sleep-deprivation.

In rat hippocampal CA1 neurons, excitatory synaptic transmission is depressed following a 20 Hz, 30 s stimulation in the stratum radiatum. This depression lasts for at least 25 min post-tetanus (long-term depression, LTD). The LTD is significantly enhanced by about 20% if rats were sleep-deprived for 12 h prior to recording. The increase in LTD following sleep-deprivation is a new finding which can have implications for hippocampal neuronal excitability, network activity, and induction of long-term potentiation and may account for post-sleep deprivation amnesia.

The involvement of GABA-C receptors in paired-pulse depression of inhibitory postsynaptic currents in rat hippocampal CA1 pyramidal neurons.

In rat hippocampal CA1 pyramidal neurons, gamma-aminobutyric acid (GABA) A receptor-mediated inhibitory postsynaptic currents (IPSCs) undergo a paired-pulse depression (PPD) by the second of two pulses, with inter-pulse intervals of 100-2000 ms, applied to the stratum radiatum. While GABA-C receptors are described in the CA1 area, their functional significance is unknown. In this study, the involvement of GABA-C receptors in PPD was examined using an in vitro hippocampal slice preparation. IPSCs evoked by stimulations in stratum radiatum were recorded with patch pipettes from CA1 pyramidal cells. PPD, when induced in the above fashion, was blocked by the GABA-C receptor antagonist (1,2,5,6-Tetrahydropyridin-4-yl) methylphosphinic acid (TPMPA, 10 muM, applied in the superfusing medium). GABA-A and GABA-B receptor-mediated IPSCs, as well as the baclofen-induced suppression of the GABA-A receptor mediated IPSC, were not antagonized by TPMPA (10-20 muM). These results indicate that PPD of the IPSC is mediated by the activation of GABA-C receptors.

A method for recording miniature inhibitory postsynaptic currents in the central nervous system suitable for quantal analysis.

Quantal analysis of transmitter release is useful in examining presynaptic mechanisms involved in synaptic transmission. However, in central neurons, the presence of multiple synapses makes it difficult to use the traditional quantal analysis, developed for the neuromuscular transmission. We developed a method to minimize these difficulties. Experiments were performed, using the whole-cell patch-clamp recording technique, on rat CA1 pyramidal neurons in a hippocampal slice preparation. When the stratum radiatum was stimulated, mixed current signals including, miniature inhibitory postsynaptic currents (mIPSCs), miniature excitatory postsynaptic currents (mEPSCs), evoked inhibitory postsynaptic currents (eIPSCs) and evoked excitatory postsynaptic currents (eEPSCs), could be observed in CA1 pyramidal cells while slices were superfused with the normal, Na(+)-containing, medium. The mIPSCs could be blocked by bicuculline (10 microm). mEPSCs, eEPSCs and eIPSCs could not be observed when the Na(+)-containing perfusion medium was replaced by a Na(+)-free medium but reappeared when the Na(+)-containing medium was re-introduced. When a polarizing electrode was placed near the recorded neuron, while slices were superfused with the Na(+)-free medium, and depolarizing rectangular current pulses of different magnitudes were applied, the number of mIPSCs increased with increasing amount of the current. Amplitudes of the mIPSCs showed a Gaussian distribution and the coefficient of variation was small. These observations indicate that a combination of the Na(+)-free superfusing medium and local depolarizations with a polarizing electrode is useful for recording mIPSCs from a localized area of the recorded neuron and for quantal analysis.

Theta-bursts induce a shift in reversal potentials for GABA-A receptor-mediated postsynaptic currents in rat hippocampal CA1 neurons.

Theta-burst stimulation of the stratum radiatum induces a negative shift in the reversal potential (RP) of gamma-aminobutyric acid (GABA)-ergic postsynaptic currents (PSCs) in hippocampal CA1 neurons in brain slices from rats of age groups 3-4 days, 6-9 days and 3-4 weeks. Furosemide reversed the shift in the RP. The amplitude of the evoked PSC appeared to increase following the theta-burst stimulation but this increase was secondary to the change in the RP. These results indicate that the RP for GABA-ergic PSCs undergoes an activity-dependent plasticity in not only neonatal but also adult neurons presumably through an up-regulation of a K(+)-Cl(-) co-transporter. This plasticity can have significant implications for neuronal network activity in the central nervous system. Furthermore, these results indicate that studies on GABA-ergic synaptic efficacy require a careful, parallel monitoring of the RP.

Theta bursts set up glutamatergic as well as GABA-ergic plasticity in neonatal rat hippocampal CA1 neurons.

gamma-Aminobutyric acid (GABA) is inhibitory in adult, but excitatory in neonatal, neurons. The switch from excitatory to inhibitory action is due to a negative shift in the equilibrium potential for the GABA(A) receptor-mediated postsynaptic current (E(GABA-PSC)). Here, we report that, in neonatal rat hippocampal CA1 neurons, presynaptic theta-burst activation induces not only a shift in E(GABA-PSC) towards that in adult neurons, but also a recruitment of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)-receptor-mediated postsynaptic currents.

Change in diazepam sensitivity of GABAA currents after LTP induction in neurons of deep cerebellar nuclei.

In deep cerebellar nuclei (DCN) neurons, inhibitory postsynaptic currents (IPSCs) undergo long-term depression (LTD) following a 10-Hz stimulation, and long-term potentiation (LTP) after a 100 Hz stimulation of the inputs. Whole-cell recordings were made from DCN neurons and changes in IPSC sensitivity to diazepam after LTD and LTP investigated. Diazepam enhanced the evoked IPSC amplitude by 45% in controls and after LTD induction. However, after LTP induction, diazepam increased the IPSC by only 16%. Diazepam increased THIP response by 34% in controls, but by only 4% after LTP. These results suggest that during LTP the diazepam sensitive GABAA receptor sub-units undergo changes.

Benzodiazepine involvement in LTP of the GABA-ergic IPSC in rat hippocampal CA1 neurons.

Benzodiazepine binding sites are present on gamma-aminobutyric acid (GABA) receptors in hippocampal neurons. Diazepam is known to potentiate the amplitude and prolong the decay of GABA(A) receptor-mediated inhibitory postsynaptic currents (IPSCs). In this study, benzodiazepine involvement in long-term potentiation (LTP) of the IPSC was examined. Whole-cell recordings of IPSCs were made from rat hippocampal CA1 neurons in a slice preparation. LTP was induced by a tetanic stimulation in the stratum radiatum (2 trains of 100 Hz for 1 s, 20 s inter-train interval) while pharmacologically blocking ionotropic glutamate receptors. During LTP, the amplitude of the IPSCs was potentiated in the majority of neurons with the IPSC decay and shape unaffected. Diazepam (5 microM) potentiated the IPSC amplitude and prolonged the decay when applied before, but not during, LTP. In neurons in which LTP could not be induced by a tetanic stimulation, diazepam did not increase the amplitude of the pre-tetanic IPSC. Flumazenil, at a concentration (10 microM) that blocked the enhancement of the IPSC by applied diazepam, had no effect on the IPSC amplitude when applied before LTP induction but significantly decreased the IPSC when applied during LTP maintenance. The antagonist, when applied during the tetanic stimulation, did not block LTP, suggesting that benzodiazepine receptors do not participate in LTP induction. These results indicate that the maintenance of LTP of the IPSC involves (a) the release of endogenous benzodiazepine agonist(s) and/or (b) the participation of benzodiazepine binding sites on subsynaptic GABA(A) receptors.

Activity-mediated shift in reversal potential of GABA-ergic synaptic currents in immature neurons.

Gamma-aminobutyric acid (GABA), which is inhibitory in the adult central nervous system, can be excitatory in the developing brain. The change from excitatory to inhibitory action of GABA during development is caused by a negative shift in its reversal potential. Here, we report a presynaptic activity-mediated negative shift in the reversal potential of the GABA-mediated synaptic currents in immature deep cerebellar nuclei neurons. This shift appears to be due to an increased expression and activation of the K+-Cl- co-transporter type 2 (KCC-2) through the activation of protein kinase A, protein synthesis and activation of protein phosphatases. Thus, maturation of the GABA response may rely on an activity-dependent up-regulation of KCC-2.

Mechanisms underlying LTP of inhibitory synaptic transmission in the deep cerebellar nuclei.

Whole-cell recordings were used to investigate long-term potentiation of inhibitory synaptic currents (IPSCs) in neurons of deep cerebellar nuclei (DCN) in slices. IPSCs were evoked by electrical stimulation of the white matter surrounding the DCN in the presence of non-N-methyl-D-aspartate (non-NMDA) glutamate receptor antagonist 6,7-dinitroquinoxaline-2,3-dione (20 microM). High-frequency stimulation induced a long-term potentation (LTP) of the IPSC amplitude without changing its reversal potential, rise time, and decay-time constant. This LTP did not require the activation of postsynaptic gamma-aminobutyric acid-A (GABA(A)) receptors but depended on the activation of NMDA receptors. LTP of IPSCs in DCN neurons could also be induced by voltage-depolarizing pulses in postsynaptic neurons and appeared to depend on an increase in intracellular calcium as the LTP was blocked when the cells were loaded with a calcium chelator, 1,2-bis-(2-amino-phenoxy)-N,N,N', N'-tetraacetic acid (BAPTA, 10 mM). LTP of IPSCs was accompanied by an increase in the frequency of spontaneous IPSCs and miniature IPSCs (recorded in the presence of tetrodotoxin 1 microM), but there was no significant change in their amplitude. In addition, during the LTP, the amplitude of response to exogenously applied GABA(A) receptor agonist 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol hydrochloride was increased. Intracellular application of tetanus toxin, a powerful blocker of exocytosis, in DCN neuron prevented the induction of LTP of IPSCs. Our results suggest that the induction of LTP of IPSCs in the DCN neurons likely involves a postsynaptic locus. Plasticity of inhibitory synaptic transmission in DCN neurons may play a crucial role in cerebellar control of motor coordination and learning.

Mechanisms involved in tetanus-induced potentiation of fast IPSCs in rat hippocampal CA1 neurons.

In the present study, possible mechanisms involved in the tetanus-induced potentiation of gamma-aminobutyric acid-A (GABA-A) receptor-mediated inhibitory postsynaptic currents (IPSCs) were investigated using the whole cell voltage-clamp technique on CA1 neurons in rat hippocampal slices. Stimulations (100 Hz) of the stratum radiatum, while voltage-clamping the membrane potential of neurons, induces a long-term potentiation (LTP) of evoked fast IPSCs while increasing the number but not the amplitude of spontaneous IPSCs (sIPSCs). The potentiation of fast IPSCs was input specific. During the period of IPSC potentiation, postsynaptic responses produced by 4,5,6,7-tetrahydroisoxazolo[5,4-c]pyridin-3-ol hydrochloride and baclofen, GABA-A and GABA-B agonists respectively, were not significantly different from control. CGP 36742, a GABA-B antagonist, blocked the induction of tetanus-induced potentiation of evoked and spontaneous IPSCs, while GTPgammaS, an activator of G proteins, substitution for GTP in the postsynaptic recording electrode did not occlude potentiation. Since GABA-B receptors work through G proteins, our results suggest that pre- but not postsynaptic GABA-B receptors are involved in the potentiation of fast IPSCs. A tetanus delivered when GABA-A responses were completely blocked by bicuculline suggests that GABA-A receptor activation during tetanus is not essential for the induction of potentiation. Rp-cAMPs, an antagonist of protein kinase A (PKA) activation, blocks the induction of potentiation of fast IPSCs. Forskolin, an activator of PKA, increases baseline evoked IPSCs as well as the number of sIPSCs, and a tetanic stimulation during this enhancement uncovers a long-term depression of the evoked IPSC. Sulfhydryl alkylating agents, N-ethylmaleimide and p-chloromercuribenzoic acid, which have been found to presynaptically increase GABA release and have been suggested to have effects on proteins involved in transmitter release processes occurring in nerve terminals, occlude tetanus-induced potentiation of evoked and spontaneous IPSCs. Taken together our results suggest that LTP of IPSCs originates from a presynaptic site and that GABA-B receptor activation, cyclic AMP/PKA activation and sulfhydryl-alkylation are involved. Plasticity of IPSCs as observed in this study would have significant implications for network behavior in the hippocampus.

Effects of hemoglobin and its breakdown products on synaptic transmission in rat hippocampal CA1 neurons.

During head injuries and hemorrhagic stroke, blood is released into the extravascular space. The pooled erythrocytes get lysed and hemoglobin is released into the intracranial cavities. Therefore, neurons may be exposed to hemoglobin and/or its breakdown products, hemin and iron, for long periods of time. In this study, the electrophysiological actions of these agents on synaptic transmission in rat hippocampal CA1 pyramidal neurons were studied using extracellular field- and whole cell patch-recordings. Previously our laboratory reported that commercially available hemoglobin produced a dose dependent suppression of synaptic transmission in hippocampal CA1 neurons. In the present study, however, we found that this depression was caused by impurities present in the hemoglobin samples. Commercially available hemoglobin and methemoglobin did not have a significant effect on synaptic transmission. Although, reduced-hemoglobin prepared using a method described by Martin et al. [J. Pharm. Exp. Ther. 232 (1985) 708], produced a significant depression of synaptic transients, these effects were due to contamination with bisulfite that was present due to the reducing procedure. Therefore, the technique of Martin et al. was inadequate in removing the reducing agents or their breakdown products. A number of studies in literature used commercial samples of hemoglobin or reduced hemoglobin prepared using the method of Martin et al. Our observations indicate that it would be important to determine if contaminants, rather than hemoglobin, are responsible for the observed effects in these studies. Unlike hemoglobin, its breakdown products, ferrous chloride and hemin, produced an irreversible and significant depression of field excitatory postsynaptic potentials. The relevance of these effects in neurological complications that follow head injuries and hemorrhagic stroke awaits further investigation.

GABA-ergic transmission in deep cerebellar nuclei.

gamma-Aminobutyric acid (GABA) is the inhibitory transmitter released at Purkinje cell axon terminals in deep cerebellar nuclei (DCN). Neurons in DCN also receive excitatory glutamatergic inputs from the inferior olive. The output of DCN neurons, which depends on the balance between excitation and inhibition on these cells, is involved in cerebellar control of motor coordination. Plasticity of synaptic transmission observed in other areas of the mammalian central nervous system (CNS) has received wide attention. If GABA-ergic and/or glutamatergic synapses in DCN also undergo plasticity, it would have major implications for cerebellar function. In this review, literature evidence for GABA-ergic synaptic transmission in DCN as well as its plasticity are discussed. Studies indicate that fast inhibitory postsynaptic potentials (IPSPs) and currents (IPSCs) in neurons of DCN are mediated by GABAA receptors. While GABAB receptors are present in DCN, they do not appear to be activated by Purkinje cell axons. The IPSPs undergo paired-pulse, as well as frequency-dependent, depressions. In addition, tetanic stimulation of inputs can induce a long-term depression (LTD) of the IPSPs and IPSCs. Excitatory synapses do not appear to undergo long-term potentiation or LTD. The LTD of the IPSP is not input-specific, as it can be induced heterosynaptically and is associated with a reduced response of DCN neurons to a GABAA receptor agonist. Postsynaptic Ca2+ and protein phosphatases appear to contribute to the LTD. The N-methyl-D-aspartate receptor-gated, as well as the voltage-gated Ca2+ channels are proposed to be sources of the Ca2+. It is suggested that LTD of GABA-ergic transmission, by regulating DCN output, can modulate cerebellar function.

Quantal release of transmitter at a central synapse.

Miniature end-plate potentials recorded at the neuromuscular junction are caused by a quantal release of acetylcholine and evoked end-plate potentials can be described as integer multiples of the miniature potentials. A variety of factors including the presence of multiple synapses on postsynaptic cells and dendritic filtering, complicate quantal analyses at central synapses. In the present investigation on rat hippocampal slices, transmitter release was blocked except for a localized area on the apical dendrites of CA1 neurons and quantal analysis was performed on miniature excitatory postsynaptic currents (mEPSCs) and evoked EPSCs of low quantum content. The results indicate that under these conditions, alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor-mediated mEPSCs exhibit a normal size distribution with S.D. values comparable to those at the neuromuscular junction, and the evoked EPSCs can be described as integer multiples of the miniature currents. The results also support reports in literature that long-term potentiation (LTP) is associated with an increase in mEPSC frequency. Whether the increase is due to (a) the enhancement of quantal release at already functional synapses, or (b) the recruitment of nearby silent synapses where a neglible transmitter release becomes measurable, or clusters of functional receptors are uncovered, cannot yet be distinguished.

Postsynaptic mechanisms underlying long-term depression of GABAergic transmission in neurons of the deep cerebellar nuclei.

1. The mechanisms underlying long-term depression (LTD) of gamma-aminobutyric acid-A (GABAA) receptor-mediated synaptic transmission induced by 10-Hz stimulation of the inhibitory afferents were investigated using perforated and whole cell voltage-clamp recordings from neurons of the deep cerebellar nuclei (DCN). 2. LTD of inhibitory postsynaptic currents (IPSCs) was reliably induced when the 10-Hz stimulation was delivered under current-clamp conditions where the postsynaptic neuronal membrane was allowed to depolarize. 3. Currents elicited by local applications of the GABAA receptor agonist, 4,5,6,7-tetrahydroisoxazolo [5,4-c]pyridin-3-ol hydrochloride (THIP) were also depressed during LTD. 4. LTD could be induced heterosynaptically and did not require the activation of GABAA receptors during the 10-Hz stimulation. 5. In cells loaded with QX-314 and superfused with media containing 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) and 2-amino-5-phosphonovaleric acid (APV), a series of depolarizing pulses (50 mV, 200 ms) induced a sustained depression of the IPSC. However, this was not observed in cells recorded with high bis-(o-aminophenoxy)-N,N,N',N'-tetraacetic acid (BAPTA)-containing pipette solutions or when they were exposed to the L-type Ca2+ channel antagonist, nitrendipine. 6. The 10-Hz-induced LTD was also inhibited by BAPTA and was significantly reduced when DCN cells were loaded with microcystin LR or treated with okadaic acid, both inhibitors of protein phosphatases. 7. These results indicate that increases in postsynaptic [Ca2+] and phosphatase activity can reduce the efficacy of GABAA receptor-mediated synaptic transmission.

Electrophysiological actions of hemoglobin on rat hippocampal CA1 pyramidal neurons.

Hemoglobin, the oxygen-carrying component of red blood cells, can be released from erythrocytes in hemorrhagic stroke and intracranial bleeding associated with head injuries. Therefore, neurons may be exposed to this agent. In addition, hemoglobin can chelate nitric oxide (NO) and has been used in studying the role of NO in synaptic plasticity and excitotoxicity. However, the electrophysiological actions of hemoglobin on central neurons are not well characterized. In the present investigation, the electrophysiological actions of hemoglobin on CA1 pyramidal neurons in rat hippocampal slices were studied with conventional intracellular pointed microelectrode- as well as perforated patch-recordings. Superfusion of rat hippocampal slices with hemoglobin (0.05 or 0.1 mM for 10-15 min) induced a depolarization of CA1 neurons and suppressed the stratum radiatum stimulation-induced excitatory postsynaptic potentials (EPSPs) and inhibitory postsynaptic potentials (IPSPs). The hemoglobin-induced depolarization as well as the suppression of the synaptic transients were present in slices pretreated with 0.1 or 0.5 mM of N omega-nitro-L-arginine, a nitric oxide synthase inhibitor, suggesting that hemoglobin has electrophysiological actions on hippocampal CA1 neurons that are independent of its NO scavenging property.

Tetanus-induced potentiation of inhibitory postsynaptic potentials in hippocampal CA1 neurons.

In the present study, tetanus-induced potentiation of inhibitory postsynaptic potentials (IPSPs), previously described by our laboratory, was further investigated in guinea pig hippocampal CA1 neurons. Tetanic stimulation of the stratum radiatum induced a long-term potentiation of the excitatory postsynaptic potential and a potentiation of the gamma-aminobutyric acid A (GABAA) receptor mediated fast IPSP without enhancing the GABAB receptor-mediated slow IPSP. During the potentiation, IPSPs evoked by stimulation of the alveus were unaffected. When slices were superfused with DL-2-amino-5-phosphonovaleric acid (an N-methyl-D-aspartate, NMDA, antagonist) and 6-cyano-7-nitroquinoxaline-2,3-dione (a non-NMDA glutamate antagonist), the potentiation of the monosynaptic fast IPSP could still be induced and maintained, suggesting that polysynaptic influences were unnecessary for this process. Finally, since the potentiation was observed in CA1 neurons in which BAPTA or K-252b was injected, this form of plasticity does not appear to be dependent on a rise in intracellular [Ca2+] or protein kinase C (PKC) activity. These results indicate that tetanic stimulation of the stratum radiatum induces a potentiation of GABAergic fast IPSPs in CA1 neurons. The potentiation may be localized to the GABAergic synapse on CA1 neurons.

Pharmacological characterization of pre- and postsynaptic GABAB receptors in the deep nuclei of rat cerebellar slices.

Whole-cell current-and voltage-clamp recordings were made from deep nuclear neurons in cerebellar slices from seven- to nine-day-old rats. Baclofen, a GABAB agonist, produced a slow postsynaptic hyperpolarization associated with a decrease in input resistance. The hyperpolarization was G-protein-dependent, blocked by intracellular Cs+ and antagonized by CGP 35348, a GABAB antagonist. In dialysed neurons recorded with Cs+ -containing pipettes, baclofen suppressed deep nuclear neuronal inhibitory postsynaptic potentials and inhibitory postsynaptic currents evoked by electrical stimulations of the Purkinje cell axons. This effect was blocked by CGP 35348, indicating that the suppressions were mediated by presynaptic GABAB receptors. The inability of CGP 35348 or uptake inhibitors (nipecotic acid and NO-711) to alter the decay of inhibitory postsynaptic currents evoked by maximal stimulation suggested that GABAB receptors are not activated by the stimulation of the GABAergic input. Paired-pulse depression of inhibitory postsynaptic currents was not blocked by CGP 35348. Moreover, neither uptake inhibitors nor CGP 35348 produced any significant changes to the whole-cell current produced by a tetanic stimulation of Purkinje cell axons, suggesting that GABAB autoreceptors were also not activated by endogenous GABA release. Our findings indicate that while pre- and postsynaptic GABAB receptors are present in the deep nuclei of the rat cerebellum, they are not activated by electrical stimulation of the Purkinje cell axons.