Katsunori Kobayashi

Impairment of hippocampal mossy fiber LTD in mice lacking mGluR2

Subtype 2 of the metabotropic glutamate receptor (mGluR2) is expressed in the presynaptic elements of hippocampal mossy fiber—CA3 synapses. Knockout mice deficient in mGluR2 showed no histological changes and no alterations in basal synaptic transmission, paired-pulse facilitation, or tetanus-induced long-term potentiation (LTP) at the mossy fiber—CA3 synapses. Long-term depression (LTD) induced by low-frequency stimulation, however, was almost fully abolished. The mutant mice performed normally in water maze learning tasks. Thus, the presynaptic mGluR2 is essential for inducing LTD at the mossy fiber—CA3 synapses, but this hippocampal LTD does not seem to be required for spatial learning.

Presynaptic long-term depression at the hippocampal Mossy fiber-CA3 synapse

Long-term potentiation (LTP) and long-term depression (LTD) of synaptic strength may underlie learning and memory in the brain. The induction of LTP occurs in postsynaptic cells in the hippocampal CA1 region but is presynaptic in CA3. LTD is also well characterized in CA1 but not in CA3. Low-frequency stimulation of mouse hippocampal slices caused homosynaptic LTD at the mossy fiber—CA3 synapse, which may be induced presynaptically by activation of metabotropic glutamate receptors. Thus, the efficacy of mossy fiber—CA3 synapses can be regulated bidirectionally, which may contribute to neuronal information processing.

Functional coupling of the nociceptin/orphanin FQ receptor with the G-protein-activated K+ (GIRK) channel

Nociceptin/orphanin FQ is a heptadecapeptide which was recently isolated from brains. It induces hyperalgesia, in contrast to the analgesic effects of opioid ligands, although it and its receptor structurally resemble opioid peptides and opioid receptors, respectively. To investigate the molecular mechanism underlying nociceptin/orphanin FQ actions, we performed Xenopus oocyte expression assays, in situ hybridization histochemistry and electrophysiological analyses of neurons. We found that the nociceptin/orphanin FQ receptor is functionally coupled with the G-protein-activated K+ (GIRK) channel in Xenopus oocytes, and that the receptor mRNA and GIRK1 mRNA co-exist in various neurons, including hippocampal pyramidal cells. Furthermore, we found that nociceptin/orphanin FQ induces hyperpolarizing currents via inward-rectifier K+ channels in hippocampal pyramidal cells, suggesting that the nociceptin/orphanin FQ receptor couples with the GIRK channel in this region. We conclude that the nociceptin/orphanin FQ receptor couples with the GIRK channel in various neurons, including hippocampal pyramidal cells, thereby modulating neuronal excitability.

Developmental Decrease in Synaptic Facilitation at the Mouse Hippocampal Mossy Fibre Synapse

Transmission at the hippocampal mossy fibre (MF)‐CA3 pyramidal cell synapse is characterized by prominent activity‐dependent facilitation, which is thought to provide a wide dynamic range in hippocampal informational flow. At this synapse in mice the magnitude of paired‐pulse facilitation and frequency‐dependent facilitation markedly decreased with postnatal development from 3 weeks (3W) to 9 weeks (9W). Throughout this period the mean amplitude and variance of unitary EPSCs stayed constant. By altering extracellular Ca2+/Mg2+ concentrations the paired‐pulse ratio could be changed to a similar extent as observed during development. However, this was accompanied by an over 30‐fold change in EPSC amplitude, suggesting that the developmental change in facilitation ratio cannot simply be explained by a change in release probability. With paired‐pulse stimulation the Ca2+ transients at MF terminals, monitored using mag‐fura‐5, showed a small facilitation, but its magnitude remained similar between 3W and 9W mice. Pharmacological tests using CNQX, adenosine, LY341495, H‐7 or KN‐62 suggested that neither presynaptic receptors (kainate, adenosine and metabotropic glutamate) nor protein kinases are responsible for the developmental change in facilitation. Nevertheless, loading the membrane‐permeable form of BAPTA attenuated the paired‐pulse facilitation in 3W mice to a much greater extent than in 9W mice, resulting in a marked reduction in age difference. These results suggest that the developmental decrease in the MF synaptic facilitation arises from a change associated with residual Ca2+, a decrease in residual Ca2+ itself or a change in Ca2+‐binding sites involved in the facilitation. A developmental decline in facilitation ratio reduces the dynamic range of MF transmission, possibly contributing to the stabilization of hippocampal circuitry.

CA2+ REGULATION IN THE PRESYNAPTIC TERMINALS OF GOLDFISH RETINAL BIPOLAR CELLS

1. To investigate regulation of the intracellular free Ca2+ concentration ([Ca2+]i) in presynaptic terminals, the Ca2+ current (ICa) and [Ca2+]i in axon terminals were simultaneously monitored in acutely dissociated retinal bipolar cells under whole‐cell voltage clamp. 2. The recovery phase of the Ca2+ transient, which was evoked by activation of ICa, became slower when the Na(+)‐Ca2+ exchanger was suppressed by removing extracellular Na+. 3. Inhibition of the plasma membrane Ca2+ pump produced by raising extracellular pH to 8.4 increased the basal [Ca2+]i and caused incomplete recovery from the Ca2+ transient. These effects were not observed in orthovanadate‐loaded bipolar cells. 4. The Ca2+ transient was not significantly affected by ryanodine, caffeine, thapsigargin, Ruthenium Red or FCCP. Internal Ca2+ stores may not participate in shaping the Ca2+ transient. 5. The ratio of the peak amplitude of the Ca2+ transient to the total amount of Ca2+ influx became smaller as the size of the Ca2+ influx increased. This action was not affected by blockage of Ca2+ transporters in the plasma membrane, or by reduction of the rate of Ca2+ influx. The peak amplitude of the Ca2+ transient seemed to be determined by Ca2+ buffering substances with a positive co‐operativity.

Calcium-dependent mechanisms involved in presynaptic long-term depression at the hippocampal mossy fibre–CA3 synapse

Long‐term potentiation (LTP) and long‐term depression (LTD) are induced presynaptically at the hippocampal mossy fibre–CA3 synapse. Activation of presynaptic metabotropic glutamate receptors (mGluRs) is necessary, but not sufficient for the LTD induction. Using mouse hippocampal slices, we attempted to identify additional presynaptic factors involved in the induction of mossy fibre LTD. Suppression of a rise in the presynaptic intracellular Ca2+ concentration ([Ca2+]i) with a membrane‐permeable Ca2+ chelator, 1,2‐bis(2‐aminophenoxy)ethane‐N,N,N′,N′‐tetraacetic acid tetraacetoxymethyl ester (BAPTA‐AM), reduced the magnitude of LTD, whereas an increase in Ca2+ influx induced LTD, suggesting that an elevation of presynaptic [Ca2+]i is crucial for the LTD induction. A broad‐spectrum protein kinase inhibitor, H‐7, blocked LTD without affecting a presynaptic inhibition induced by an mGluR agonist. Furthermore, LTD was reduced by an inhibitor of calmodulin or Ca2+/calmodulin‐dependent protein kinases. Thus, we conclude that mossy fibre LTD requires an increase in presynaptic [Ca2+]i and subsequent activation of Ca2+/calmodulin‐dependent protein kinases. Because mossy fibre LTP may also require a rise in presynaptic [Ca2+]i, bidirectional long‐term plasticity at the mossy fibre synapse is likely to be regulated by presynaptic Ca2+‐dependent processes.

Platelet‐activating factor receptor is not required for long‐term potentiation in the hippocampal CA1 region

From pharmacological studies, platelet‐activating factor (PAF) has been proposed as a retrograde messenger for long‐term potentiation (LTP) in the hippocampal CA1 region. We re‐examined a possible contribution of PAF to LTP with a more specific approach using mice deficient in the PAF receptor. The PAF receptor‐deficient mice exhibited normal LTP and showed no obvious abnormality in excitatory synaptic transmission. We also performed pharmacological experiments on the wild‐type mice. Two structurally different antagonists of PAF receptors had no effects on LTP. Furthermore, the application of PAF itself caused no detectable changes in excitatory synaptic transmission. Thus, we conclude that the PAF receptor is not required for LTP in the CA1 region.

Potentiation of Ca2+ transients in the presynaptic terminals of goldfish retinal bipolar cells.

1. To study a possible contribution of intracellular Ca2+ stores to the presynaptic Ca2+ regulation, the Ca2+ current (ICa) and the intracellular free Ca2+ concentration ([Ca2+]i) were simultaneously monitored in isolated goldfish retinal bipolar cells using the whole‐cell voltage clamp procedure and fura‐2 fluorimetry. 2. The Ca2+ transient triggered by the activation of ICa was potentiated when [Ca2+]i was increased by applying either a prepulse or a small steady depolarization. The potentiation seemed to be partly due to the release of Ca2+ from intracellular Ca2+ stores. 3. The intracellular Ca2+ release was reversibly inhibited by caffeine but was not affected by ryanodine, suggesting that Ca2+ is released through intracellular Ca2+ channels which differ from ryanodine receptor channels. 4. These results suggest that the intracellular Ca2+ release may contribute to the facilitation of transmitter release.