Metabotropic glutamate receptor-mediated depression of the slow afterhyperpolarization is gated by tyrosine phosphatases in hippocampal CA1 pyramidal neurons

Group I metabotropic glutamate receptor (mGluR) agonists increase the excitability of hippocampal CAl pyramidal neurons via depression of the postspike afterhyperpolarization. In adult rats, this is mediated by both mGluR1 and -5, but the signal transduction processes involved are unknown. In this study, we investigated whether altered levels of tyrosine phosphorylation of proteins are involved in the depression of the slow-duration afterhyperpolarization (sAHP) by the Group I mGluR agonist (RS)-3,5-dihydroxyphenylglycine (DHPG) in CA1 pyramidal neurons of rat hippocampal slices. Preincubation with the tyrosine kinase inhibitors lavendustin A or genistein, or the Src-specific inhibitor 3-(4-chlorophenyl) 1-(1,1-dimethylethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-amine (PP2), did not inhibit the DHPG-mediated depression of the sAHP. However, preincubation with the tyrosine phosphatase inhibitor orthovanadate reduced the effects of DHPG. This effect of orthovanadate was prevented by simultaneous inhibition of tyrosine kinases with lavendustin A. Selective activation of either mGluR1 or -5 by application of DHPG plus either the mGluR5 antagonist 2-methyl-6-(phenylethynyl)pyridine (MPEP) or the mGluR1 antagonist (S)-(+)-alpha-amino-4-carboxy-2-methylbenzeneacetic acid (LY367385) demonstrated that the effect of inhibiting tyrosine phosphatases is not specific to either subtype of mGluR. These results suggest that the depression of the sAHP induced by activation of mGluR1 and -5 is gated by a balance between tyrosine phosphorylation and dephosphorylation.     

mGluR1, but not mGluR5, mediates depolarization of spinal cord neurons by blocking a leak current

The modulation of neuronal excitability by group I metabotropic glutamate receptors (mGluRs) was studied in isolated lamprey spinal cord. At resting potential, application of the group I mGluR agonist (R,S)-3,5-dihydroxyphenylglycine (DHPG) slightly depolarized the cells. However, at depolarized membrane potentials, this agonist induced repetitive firing. When Na+ channels were blocked by TTX, DHPG induced a slight depolarization at rest that increased in amplitude as the neurons were held at more depolarized membrane potentials. In voltage-clamp conditions, DHPG application induced an inward current associated with a decrease in membrane conductance when cells were held at -40 mV. At resting membrane potential, no significant change in the current was induced by DHPG, although a decrease in membrane conductance was seen. The conductance blocked by DHPG corresponded to a leak current, since DHPG had no effect on the voltage-gated current elicited by a voltage step from -60 to -40 mV, when leak currents were subtracted. The leak current blocked by DHPG is mediated by fluxes of both K+ and Na+. The subtype of group I mGluR mediating the block of the leak current was characterized using specific antagonists for mGluR1 and mGluR5. The inhibition of the leak current was blocked by the mGluR1 antagonist LY 367385 but not by the mGluR5 antagonist 2-methyl-6-(phenylethynyl)pyridine (MPEP). The DHPG-induced blockage of the leak current required phospholipase C (PLC)-activation and release of Ca2+ from internal stores as the effect of DHPG was suppressed by the PLC-blocker U-73122 and after depletion of intracellular Ca2+ pools by thapsigargin. Our results thus show that mGluR1 activation depolarizes spinal neurons by inhibiting a leak current. This will boost membrane depolarization and result in an increase in the excitability of spinal cord neurons, which could contribute to the modulation of the activity of the spinal locomotor network.     

Group I metabotropic glutamate receptors on second-order baroreceptor neurons are tonically activated and induce a Na+-Ca2+ exchange current

The nucleus tractus solitarius (NTS) is essential for coordinating baroreflex control of blood pressure. The baroreceptor sensory fibers make glutamatergic synapses onto second-order NTS neurons. Glutamate spillover activates Group II and III presynaptic metabotropic glutamate receptors (mGluRs) on the baroreceptor central terminals to inhibit synaptic transmission, but the role of postsynaptic mGluRs is less understood. We used whole cell patch-clamping in anatomically identified second-order baroreceptor neurons in a brain stem slice to test whether Group I, II, and III mGluRs had postsynaptic effects at this first central synapse in the baroreceptor afferent pathway. The Group I agonist DHPG induced a depolarization and spiking that was mimicked by endogenous glutamate. Group I mGluR blockade prevented the depolarization and slightly hyperpolarized the neurons, suggesting a small tonic Group I mGluR activation. The DHPG-induced inward current consisted of voltage-dependent and -independent components; the former was blocked by TEA and the latter was blocked by replacing extracellular NaCl with LiCl or Tris-HCl. The DHPG current was potentiated in a Ca2+-free external solution and was diminished by intracellular dialysis with BAPTA and by perfusion with Na+-Ca2+ exchanger blockers, KB-R7943 or 3',4'-dichlorobenzamil. Intracellular dialysis with GDPbetaS or heparin and perfusion with the PLC inhibitor U-73122 or the Ca2+-calmodulin inhibitor W-7 significantly decreased the DHPG current. The data suggest that Group I mGluRs on baroreceptor neurons are functional; are activated by endogenous glutamate; and activate a Na+-Ca2+ exchanger through G-protein, PLC, IP3, and Ca2+-calmodulin mechanisms to excite the cell, thus providing postsynaptic mechanisms to enhance or prolong baroreceptor signal transmission.     

Activation of group I metabotropic glutamate receptors leads to brain-derived neurotrophic factor expression in rat C6 cells

Brain-derived neurotrophic factor (BDNF), which mediates neuronal growth, neuroprotection and synaptic modulation, is expressed in neurons and glial cells. The present study investigated the expression of BDNF in response to the activation of group I metabotropic glutamate receptors (mGluRs) by (S)-3,5-Dihydroxyphenylglycine (DHPG) in rat C6 glioma cells. The increase in BDNF mRNA in DHPG-stimulated cells, which peaked by 12 h after DHPG exposure, was attenuated by the mGluR5 inhibitor MPEP, but not by the mGluR1 inhibitor CPCCOEt. DHPG-induced BDNF mRNA expression reduced in cultures pretreated with protein kinase C (PKC) inhibitor, GFX, but not with calcium/calmodulin kinase II (CaMKII) inhibitor, KN-93. Immunostaining revealed high BDNF expression in cytoplasm of C6 cells after 48 h of incubation with 1 μM DHPG, but this was lower in MPEP-pretreated cells. These results indicate that activation of group I mGluRs induces BDNF mRNA and protein expression via mGluR5 subtype and PKC-dependent signaling pathway in C6 glioma cells.     

Modulation of afterpotentials and firing pattern in guinea pig CA3 neurones by group I metabotropic glutamate receptors

Activation of group I metabotropic glutamate receptors (mGluRs) alters the firing patterns of individual CA3 pyramidal cells in guinea pig hippocampal slices. Following addition of the selective group I agonist (S)-3,5-dihydroxyphenylglycine (DHPG) to the bathing solution, pyramidal cells initially firing regular, single action potentials switched to firing in brief bursts. This change in firing pattern resulted from modulation by mGluRs of three afterpotentials. The medium and slow afterhyperpolarizations (m and sAHPs) were blocked by mGluR activation. In addition, a voltage-dependent afterdepolarization (ADP) was induced. Recordings from mutant mice lacking phospholipase C_β1 (PLC_β1) showed that mGluR block of the mAHP, as well as induction of the ADP, depended on the phosphoinositide hydrolysis pathway. Block of the sAHP, however, was partly spared in the absence of PLC_β1. Optical recordings of postspike intracellular Ca²⁺ rises showed that mGluR block of the AHP was not mediated by alterations of action potential-associated Ca²⁺ increases (Ca²⁺ transients). The mGluR induction of an ADP was also independent of any changes in the Ca²⁺ transient. The mGluR-induced change in the firing pattern of hippocampal pyramidal cells is thus the result of multiple mechanisms, including suppression of both m and sAHPs and activation of an ADP, that act together to produce a specific excitatory effect, namely an increased likelihood that a single action potential will lead immediately to one or more following action potentials.     

Signaling mechanisms underlying group I mGluR-induced persistent AHP suppression in CA3 hippocampal neurons

Activation of group I metabotropic glutamate receptors (mGluRs) leads to a concerted modulation of spike afterpotentials in guinea pig hippocampal neurons including a suppression of both medium and slow afterhyperpolarizations (AHPs). Suppression of AHPs may be long-lasting, in that it persists after washout of the agonist. Here, we show that persistent AHP suppression differs from short-term, transient suppression in that distinct and additional signaling processes are required to render the suppression persistent. Persistent AHP suppression followed DHPG application for 30 min, but not DHPG application for 5 min. Persistent AHP suppression was temperature dependent, occurring at 30–31°C, but not at 25–26°C. Preincubation of slices in inhibitors of protein synthesis (cycloheximide or anisomycin) prevented the persistent suppression of AHPs by DHPG. Similarly, preincubation of slices in an inhibitor of p38 MAP kinase (SB 203580) prevented persistent AHP suppression. In contrast, a blocker of p42/44 MAP kinase activation (PD 98059) had no effect on persistent AHP suppression. Additionally, we show that the mGluR5 antagonist MPEP, but not the mGluR1 antagonist LY 367385, prevented DHPG-induced persistent AHP suppression. Thus persistent AHP suppression by DHPG in hippocampal neurons requires activation of mGluR5. In addition, activation of p38 MAP kinase signaling and protein synthesis are required to impart persistence to the DHPG-activated AHP suppression.     

Capsaicin prevents the hyperalgesia induced by peripheral group I mGluRs activation

Group 1 metabotropic glutamate receptors (mGluRs) are expressed in peripheral and central neural tissues and involved in peripheral and central sensitization in various pain models. However, there are limited reports that activation of peripheral group I mGluRs could evoke pain. Furthermore, any behavioral evidences could not be found out, showing what kind of afferent fibers are involved in peripheral mGluRs-mediated hyperalgesia. This study was undertaken to clarify whether peripherally injected group I mGluRs agonists could induce pain-related behaviors and capsaicin-sensitive afferent fibers might be involved in the hyperalgesia. To assess pain sensitivity, mechanical threshold for paw withdrawal response (PWT) was measured and number of spontaneous flinching behavior was counted. Intraplantar injection of group I mGluR agonist, (RS)-3,5-dihydroxyphenylglycine (DHPG) and mGluR5 agonist, (RS)-2-chloro-5-hydroxyphenyglycine (CHPG) immediately induced pain-like behaviors, such as decrease of PWT and increased number of flinchings. These agonists-induced pain-like behaviors were blocked by group I mGluRs antagonist, (RS)-1-aminoindan-1,5-dicarboxylic acid (AIDA) and mGluR5 antagonist, 2-methyl-6-(phenylethynyl) pyridine hydrochloride (MPEP). Perineural pretreatment of 1% capsaicin solution significantly reduced pain-related behaviors induced by DHPG and CHPG, proposing that capsaicin-sensitive primary afferent fibers could be responsible for the hyperalgesia induced by activation of peripheral group I mGluRs. This study presents the first behavioral evidence that peripheral group I mGluRs activation could induce spontaneous as well as mechanical hyperalgesia and capsaicin-sensitive afferent fiber could be implicated the group I mGluR mediated hyperalgesia.     

Regulation of phosphorylation of NMDA receptor NR1 subunits in the rat neostriatum by group I metabotropic glutamate receptors in vivo

Group I metabotropic glutamate receptors (mGluRs) are Gαq-protein-coupled receptors and are densely expressed in medium-sized spiny projection neurons of the neostriatum. Among different subtypes of glutamate receptors, group I mGluRs have been demonstrated to actively interact with the ionotropic glutamate receptor N-methyl-d-aspartate (NMDA) subtypes for regulating various forms of cellular activities and synaptic plasticity. In this study, the possible role of group I mGluRs in regulating serine phosphorylation of NMDA receptor NR1 subunits in the neostriatum was investigated in vivo. We found in chronically cannulated rats that injection of the group I mGluR agonist 3,5-dihydroxyphenylglycine (DHPG) into the dorsal striatum (caudate putamen) significantly increased phosphorylation of the two serine residues (serine 896 and serine 897) on the intracellular C-terminus of the NR1. The increase in NR1 phosphorylation was dose-dependent and DHPG had no effect on basal levels of NR1 proteins. Intrastriatal infusion of the group I mGluR antagonist N-phenyl-7-(hydroxyimino)cyclopropa[b]chromen-1a-carboxamide (PHCCC) significantly attenuated the DHPG-stimulated NR1 phosphorylation. Pretreatment with the mGluR5 antagonist 2-methyl-6-(phenylethynyl)pyridine hydrochloride (MPEP) also produced the same effect. These data suggest that group I mGluRs, likely mGluR5 subtypes, possess the ability to upregulate protein phosphorylation of NMDA receptor NR1 subunits in striatal neurons in vivo.     

Postsynaptic induction and presynaptic expression of group 1 mGluR-dependent LTD in the hippocampal CA1 region

Activation of metabotropic glutamate receptors (mGluRs) with the group I mGluR selective agonist (R,S)-3,5-dihydroxyphenylglycine (DHPG) induces a long-term depression (LTD) of excitatory synaptic transmission in the CA1 region of the hippocampus. Here we investigated the potential roles of pre- and postsynaptic processes in the DHPG-induced LTD at excitatory synapses onto hippocampal pyramidal cells in the mouse hippocampus. Activation of mGluRs with DHPG, but not ACPD, induced LTD at both Schaffer collateral/commissural fiber synapses onto CA1 pyramidal cells and at associational/commissural fiber synapses onto CA3 pyramidal cells. DHPG-induced LTD was blocked when the G-protein inhibitor guanosine-5'-O-(2-thiodiphosphate) was selectively delivered into postsynaptic CA1 pyramidal cells via an intracellular recording electrode, suggesting that DHPG depresses synaptic transmission through a postsynaptic, GTP-dependent signaling pathway. The effects of DHPG were also strongly modulated, however, by experimental manipulations that altered presynaptic calcium influx. In these experiments, we found that elevating extracellular Ca(2+) concentrations ([Ca(2+)](o)) to 6 mM almost completely blocked the effects of DHPG, whereas lowering [Ca(2+)](o) to 1 mM significantly enhanced the ability of DHPG to depress synaptic transmission. Enhancing Ca(2+) influx by prolonging action potential duration with bath applications of the K(+) channel blocker 4-aminopyridine (4-AP) also strongly reduced the effects of DHPG in the presence of normal [Ca(2+)](o) (2 mM). Although these findings indicate that alterations in Ca(2+)-dependent signaling processes strongly regulate the effects of DHPG on synaptic transmission, they do not distinguish between potential pre- versus postsynaptic sites of action. We found, however, that while inhibiting both pre- and postsynaptic K(+) channels with bath-applied 4-AP blocked the effects of DHPG; inhibition of postsynaptic K(+) channels alone with intracellular Cs(+) and TEA had no effect on the ability of DHPG to inhibit synaptic transmission. This suggests that presynaptic changes in transmitter release contribute to the depression of synaptic transmission by DHPG. Consistent with this, DHPG induced a persistent depression of both AMPA and N-methyl-D-aspartate receptor-mediated components of excitatory postsynaptic currents in voltage-clamped pyramidal cells. Together our results suggest that activation of postsynaptic mGluRs suppresses transmission at excitatory synapses onto CA1 pyramidal cells through presynaptic effects on transmitter release.     

Peripheral metabotropic glutamate receptor 5 mediates mechanical hypersensitivity in craniofacial muscle via protein kinase C dependent mechanisms

We previously demonstrated that peripherally located N-methyl-D-aspartic acid (NMDA) receptors contribute to acute muscle nociception and the development of chronic muscular hyperalgesia. In the present study, we investigated the potential role of peripheral group I metabotropic glutamate receptors (mGluRs 1/5) in the development of muscular hypersensitivity to mechanical stimulation, and attempted to elucidate intracellular signaling mechanisms associated with the mGluR activation in male Sprague-Dawley rats. First, our Western blot analyses revealed that mGluR 5 protein, but not mGluR 1 protein, is reliably detected in trigeminal ganglia and the masseter nerve. Subsequent behavioral studies demonstrated that the group I mGluR agonist, R,S-3,5-dihydroxyphenylglycol (DHPG), significantly decreased the mechanical threshold to noxious stimulation of the masseter, and that the DHPG-induced mechanical hypersensitivity can be effectively prevented by pretreatment of the masseter with 2-methyl-6-(phenylethynyl)pyridine hydrochloride (MPEP), a selective mGluR 5 antagonist, but not by 7-(hydroxyimino)cyclopropa[b]chromen-1a-carboxylate ethyl ester (CPCCOEt), a selective mGluR 1 antagonist. Moreover, the DHPG-induced mechanical hypersensitivity was significantly blocked by inhibiting either the alpha or epsilon isoform of protein kinase C (PKC). Collectively, these data provide evidence that peripherally located mGluR 5 may play an important role in the development of masseter hypersensitivity, and that PKC activation is required for the modulatory effect of peripheral mGluR 5 in the craniofacial muscle tissue. Thus, selective targeting of peripheral mGluR 5 and PKCalpha, as well as PKCepsilon, might serve as an effective therapeutic strategy in the management of chronic muscle pain conditions, such as temporomandibular disorders.     

Activation of group I mGluRs elicits different responses in murine CA1 and CA3 pyramidal cells

The group I metabotropic glutamate receptor agonist DHPG has been shown to produce two major effects on CA3 pyramidal cells at rest: a reduction in the background conductance and an activation of a voltage-gated inward current ( I _mGluR(V)). Both effects contribute to depolarising CA3 pyramidal cells and the latter has been implicated in eliciting prolonged epileptiform population bursts. We observed that DHPG-induced depolarisation was smaller in CA1 pyramidal cells than in CA3 cells. Voltage clamp studies revealed that while DHPG elicited I _mGluR(V) in CA3 pyramidal cells, such a response was absent in CA1 pyramidal cells. Both mGluR1 and mGluR5 have been localised in CA3 pyramidal cells, whereas only mGluR5 has been detected in CA1 pyramidal cells. Using mGluR1 knockout mice, we evaluated whether the absence of an I _mGluR(V) response can be correlated with the absence of mGluR1. In these experiments, DHPG failed to elicit I _mGluR(V) in CA3 pyramidal cells. This suggests that the smaller depolarising effects of DHPG on wild-type CA1 pyramidal cells is caused, at least in part, by the absence of I _mGluR(V) in these cells and that the difference in the responses of CA1 and CA3 cells may be attributable to the lack of mGluR1 in CA1 pyramidal cells.     

Involvement of metabotropic glutamate receptor 5 in cardiomyocyte differentiation from mouse embryonic stem cells

Metabotropic glutamate receptors (mGluRs) are G-protein coupled receptors (GPCRs) activated by glutamate. The function of mGluRs is not restricted to the regulation of synaptic transmission. Although some roles of mGluR5 in mouse embryonic stem cells (ESCs) have been proposed, little is known about the significance of mGluR5 in cardiomyocyte differentiation from ESCs. We demonstrated that mGluR5 expression increased during cardiomyocyte differentiation. Activation of mGluR5 with (RS)-3, 5-dihydroxy phenylglycine (DHPG) promoted cardiomyocyte differentiation in a dose-dependent manner. DHPG significantly enhanced PI 3-kinase enhancer (PIKE) and PI3K p110α expression, but had no significant effect on Homer1b/c. The coexpression of PIKE or PI3K p110α together with Troponin T in embryoid bodies (EBs) treated with DHPG was elevated to 9.51% and 12.05%, respectively. Inhibition of mGluR5 with 2-methyl-6-(phenylethynyl)pyridine (MPEP) treating the ESCs, did hold back the cardiogenesis from the ESCs at the early differentiation stage. However, EBs applied by MPEP could not inhibit cardiomyocyte differentiation. Small interfering RNA (siRNA) of mGluR5 blocked cardiomyocyte differentiation by repressing PIKE and PI3K p110α expression, but had no notable influence on Homer1b/c. mGluR5 siRNA also decreased the DHPG-induced Ca2? transient peak amplitude in the isolated ESC-derived cardiomyocytes. The amplitude of Ca2? oscillation was reduced by ～90% with si-mGluR5-3 compared with si-control. The protein expression of T-type Ca2? channel and L-type Ca2? channel was decreased in si-mGluR5-3-treated EBs. Taken together, these results revealed that mGluR5/PIKE/PI3K signaling pathway was involved in cardiomyocyte differentiation from ESCs. The key function of mGluR5 is probably associated with cardiogenesis and Ca2? signal in ESC-derived cardiomyocytes.     

大鼠纹状体I组代谢型谷氨酸受体激动引起动物向对侧旋转

目的:研究代谢型谷氨酸受体(mGluRs)激动剂引起大鼠向对侧旋转时介导的受体亚型。方法:大鼠纹状体内微量注射mGluRs激动剂或拮抗剂,观察大鼠的意识、行为变化,并于给药后6h测定旋转活动。结果:mGluRs非亚型特异的激动剂tACPD(500、1000nmol)纹状体内注射引起大鼠向对侧旋转,mGluRs的非竞争性拮抗剂L-AP₃、竞争性拮抗剂MCPG及抑制细胞内钙释放的胆罗啉均可减轻tACPD引起的旋转。I组mGluRs的特异性激动剂DHPG(500nmol)纹状体内注射也引起大鼠向对侧旋转,MCPG及mGluR₁的拮抗剂LY367385及mGluR₅的拮抗剂MPEP均可拮抗DHPG引起的旋转。预先腹腔注射利血平(5mg/kg)可阻断DHPG的作用。结论:I组mGluRs激动引起大鼠向对侧旋转,此作用可能与细胞内钙释放有关及依赖于多巴胺的存在。     

beta-Amyloid peptides impair PKC-dependent functions of metabotropic glutamate receptors in prefrontal cortical neurons

The metabotropic glutamate receptors (mGluRs) have been implicated in cognition, memory, and some neurodegenerative disorders, including the Alzheimer's disease (AD). To understand how the dysfunction of mGluRs contributes to the pathophysiology of AD, we examined the beta-amyloid peptide (Abeta)-induced alterations in the physiological functions of mGluRs in prefrontal cortical pyramidal neurons. Two potential targets of mGluR signaling involved in cognition, the GABAergic system and the N-methyl-d-aspartate (NMDA) receptor, were examined. Activation of group I mGluRs with (S)-3,5-dihydroxyphenylglycine (DHPG) significantly increased the spontaneous inhibitory postsynaptic current (sIPSC) amplitude, and this effect was protein kinase C (PKC) sensitive. Treatment with Abeta abolished the DHPG-induced enhancement of sIPSC amplitude. On the other hand, activation of group II mGluRs with (2R,4R)-4-aminopyrrolidine-2,4-dicarboxylate (APDC) significantly increased the NMDA receptor (NMDAR)-mediated currents via a PKC-dependent mechanism, and Abeta treatment also diminished the APDC-induced potentiation of NMDAR currents. In Abeta-treated slices, both DHPG and APDC failed to activate PKC. These results indicate that the mGluR regulation of GABA transmission and NMDAR currents is impaired by Abeta treatment probably due to the Abeta-mediated interference of mGluR activation of PKC. This study provides a framework within which the role of mGluRs in normal cognitive functions and AD can be better understood.     

Group I metabotropic glutamate receptors (mGluRs) modulate visual responses in the superficial superior colliculus of the rat

Group I metabotropic glutamate receptors (mGluRs) are expressed in cells in the superficial layers of the rat superior colliculus (SSC) and SSC afferents. The purpose of this study was to investigate the physiological effect of Group I mGluR activation on visual responses of SSC neurones using both in vivo and in vitro techniques. In the in vivo preparation, agonists and antagonists were applied by iontophoresis and single neurone activity was recorded extracellularly in anaesthetised rats. Application of the Group I agonist ( S )-3,5-dihydroxyphenylglycine (DHPG) resulted in a reversible inhibition of the visual response. The effect of DHPG could be blocked by concurrent application of the Group I (mGluR1/mGluR5) antagonist ( S )-4-carboxyphenylglycine (4CPG) or mGluR1 antagonist (+)-2-methyl-4-carboxyphenylglycine (LY367385). Application of 4CPG alone resulted in a facilitation of the visual response and this effect was not changed when the visual stimulus contrast was varied. Response habituation was observed when visual stimuli were presented at 0.5 s intervals, but this was not affected by DHPG or 4CPG. In slices of the superior colliculus, stimulation of the optic tract resulted in a field EPSP recorded from the SSC whose duration was increased in the presence of the GABA antagonists picrotoxin and CGP55845. Application of DHPG (5-100 μM) reduced the field EPSP, and this effect could be reversed by the mGluR1 antagonist LY367385 (200 μM), but not by the mGluR5 antagonist MPEP (5 μM). These data show that activation of mGluR1, but probably not mGluR5, can modulate visual responses of SSC neurones in vivo , and that this could be via presynaptic inhibition of glutamate release from either retinal or, possibly, cortical afferents.     

Differential roles for mGluR1 and mGluR5 in the persistent prolongation of epileptiform bursts.

Transient activation of group I metabotropic glutamate receptors (mGluRs) with the selective agonist (S)-3,5-dihydroxyphenylglycine (DHPG) produces persistent prolongation of epileptiform bursts in guinea pig hippocampal slices, the maintenance of which can be reversibly suppressed with group I mGluR antagonists. To determine the relative roles of mGluR1 and mGluR5 in these group I mGluR-dependent induction and maintenance processes, subtype-selective antagonists were utilized. In the presence of picrotoxin, DHPG (50 microM, 20-45 min) converted interictal bursts into 1- to 3-s discharges that persisted for hours following washout of the mGluR agonist. 2-methyl-6-(phenylethynyl)-pyridine (MPEP, an mGluR5 antagonist; 25 microM) and (+)-2-methyl-4-carboxyphenylglycine (LY367385, an mGluR1 antagonist; 20-25 microM) each significantly suppressed the ongoing expression of the mGluR-induced prolonged bursts. However, LY367385 was more effective, reducing the burst prolongation by nearly 90%; MPEP only produced a 64% reduction in burst prolongation. Nevertheless, MPEP was more effective at preventing the induction of the burst prolongation; all 10 slices tested failed to express prolonged bursts both during and after co-application of DHPG with MPEP. Co-application of DHPG with LY367385, in contrast, resulted in significant burst prolongation (in 68% of slices tested) that was revealed on washout of the two agents. These results suggest that while both receptor subtypes participate in both the induction and maintenance of mGluR-mediated burst prolongation, mGluR1 activation plays a greater role in sustaining the expression of prolonged bursts, whereas mGluR5 activation may be a more critical contributor to the induction process underlying this type of epileptogenesis.     

Regulation of main olfactory bulb mitral cell excitability by metabotropic glutamate receptor mGluR1

In the rodent main olfactory bulb (MOB), mitral cells (MCs) express high levels of the group I metabotropic glutamate receptor (mGluR) subtype, mGluR1. The significance of this receptor in modulating MC excitability is unknown. We investigated the physiological role of mGluR1 in regulating MC activity in rat and mouse MOB slices. The selective group I agonist (RS)-3,5-dihydroxyphenylglycine (DHPG), but not group II or III agonists, induced potent, dose-dependent, and reversible depolarization and increased firing of MCs. These effects persisted in the presence of blockers of fast synaptic transmission, indicating that they are due to direct activation of mGluRs on MCs. Voltage-clamp recordings showed that DHPG elicited a voltage-dependent inward current consisting of multiple components sensitive to potassium and calcium channel blockade and intracellular calcium chelation. MC excitatory responses to DHPG were absent in mGluR1 knockout mice but persisted in mGluR5 knockout mice. Broad-spectrum LY341495, MCPG, as well as preferential mGluR1 LY367385 antagonists blocked the excitatory effects of DHPG and also potently modulated MC spontaneous and olfactory nerve-evoked excitability. mGluR antagonists altered spontaneous membrane potential bistability, increasing the duration of the up and down states. mGluR antagonists also substantially attenuated MC responses to sensory input, decreasing the probability and increasing the latency of olfactory nerve-evoked spikes. These findings suggest that endogenous glutamate tonically modulates MC excitability and responsiveness to olfactory nerve input, and hence the operation of the MOB circuitry, via activation of mGluR1.     

Calcium-activated afterhyperpolarizations regulate synchronization and timing of epileptiform bursts in hippocampal CA3 pyramidal neurons

Calcium-activated potassium conductances regulate neuronal excitability, but their role in epileptogenesis remains elusive. We investigated in rat CA3 pyramidal neurons the contribution of the Ca(2+)-activated K(+)-mediated afterhyperpolarizations (AHPs) in the genesis and regulation of epileptiform activity induced in vitro by 4-aminopyridine (4-AP) in Mg(2+)-free Ringer. Recurring spike bursts terminated by prolonged AHPs were generated. Burst synchronization between CA3 pyramidal neurons in paired recordings typified this interictal-like activity. A downregulation of the medium afterhyperpolarization (mAHP) paralleled the emergence of the interictal-like activity. When the mAHP was reduced or enhanced by apamin and EBIO bursts induced by 4-AP were increased or blocked, respectively. Inhibition of the slow afterhyperpolarization (sAHP) with carbachol, t-ACPD, or isoproterenol increased bursting frequency and disrupted burst regularity and synchronization between pyramidal neuron pairs. In contrast, enhancing the sAHP by intracellular dialysis with KMeSO(4) reduced burst frequency. Block of GABA(A-B) inhibitions did not modify the abnormal activity. We describe novel cellular mechanisms where 1) the inhibition of the mAHP plays an essential role in the genesis and regulation of the bursting activity by reducing negative feedback, 2) the sAHP sets the interburst interval by decreasing excitability, and 3) bursting was synchronized by excitatory synaptic interactions that increased in advance and during bursts and decreased throughout the subsequent sAHP. These cellular mechanisms are active in the CA3 region, where epileptiform activity is initiated, and cooperatively regulate the timing of the synchronized rhythmic interictal-like network activity.     

Differential roles for group 1 mGluR subtypes in induction and expression of chemically induced hippocampal long-term depression

Although metabotropic glutamate receptors (mGluRs) mGluR1 and mGluR5 are often found to have similar functions, there is considerable evidence that the two receptors also serve distinct functions in neurons. In hippocampal area CA1, mGluR5 has been most strongly implicated in long-term synaptic depression (LTD), whereas mGluR1 has been thought to have little or no role. Here we show that simultaneous pharmacological blockade of mGluR1 and mGluR5 is required to block induction of LTD by the group 1 mGluR agonist, (RS)-3,5-dihydroxyphenylglycine (DHPG). Blockade of mGluR1 or mGluR5 alone has no effect on LTD induction, suggesting that activation of either receptor can fully induce LTD. Consistent with this conclusion, mGluR1 and mGluR5 both contribute to activation of extracellular signal-regulated kinase (ERK), which has previously been shown to be required for LTD induction. In contrast, selective blockade of mGluR1, but not mGluR5, reduces the expression of LTD and the associated decreases in AMPA surface expression. LTD is also reduced in mGluR1 knockout mice confirming the involvement of mGluR1. This shows a novel role for mGluR1 in long-term synaptic plasticity in CA1 pyramidal neurons. In contrast to DHPG-induced LTD, synaptically induced LTD with paired-pulse low-frequency stimulation persists in the pharmacological blockade of group 1 mGluRs and in mGluR1 or mGluR5 knockout mice. This suggests different receptors and/or upstream mechanisms for chemically and synaptically induced LTD.     

Endocannabinoid- and mGluR5-dependent short-term synaptic depression in an isolated neuron/bouton preparation from the hippocampal CA1 region

Endocannabinoids released from the postsynaptic neuronal membrane can activate presynaptic CB1 receptors and inhibit neurotransmitter release. In hippocampal slices, depolarization of the CA1 pyramidal neurons elicits an endocannabinoid-mediated inhibition of gamma-aminobutyric acid release known as depolarization-induced suppression of inhibition (DSI). Using the highly reduced neuron/synaptic bouton preparation from the CA1 region of hippocampus, we have begun to examine endocannabinoid-dependent short-term depression (STD) of inhibitory synaptic transmission under well-controlled physiological and pharmacological conditions in an environment free of other cells. Application of the CB1 synthetic agonist WIN55212-2 and endogenous cannabinoids 2-AG and anandamide produced a decrease in spontaneous inhibitory postsynaptic current (sIPSC) frequency and amplitude, indicating the presence of CB1 receptors at synapses in this preparation. Endocannabinoid-dependent STD is different from DSI found in hippocampal slices and the neuron/bouton preparation from basolateral amygdala (BLA) since depolarization alone was not sufficient to induce suppression of sIPSCs. However, concurrent application of the metabotropic glutamate receptor (mGluR) agonist (RS)-3,5-dihydroxyphenylglycine (DHPG) and postsynaptic depolarization resulted in a transient (30-50 s) decrease in sIPSC frequency and amplitude. Application of DHPG alone had no effect on sIPSCs. The depolarization/DHPG-induced STD was blocked by the CB1 antagonist SR141716A and the mGluR5 antagonist MPEP and was sensitive to intracellular calcium concentration. Comparing the present findings with earlier work in hippocampal slices and BLA, it appears that endocannabinoid release is less robust in isolated hippocampal neurons.