Modulating mechanosensory afferent excitability by an atypical mGluR

Mechanotransduction by proprioceptive sensory organs is poorly understood. Evidence was recently shown that muscle spindle and hair follicle primary afferents (lanceolates) constantly release glutamate from synaptic‐like vesicles (SLVs) within the terminals. The secreted glutamate activates a highly unusual metabotropic glutamate receptor (mGluR) to modulate the firing rate (spindles) and SLV recycling (lanceolates). This receptor has yet to be isolated and sequenced. To further investigate this receptor's pharmacology, ligands selective for classical mGluRs have been recently characterised for their ability to alter stretch‐evoked spindle firing and SLV endocytosis in these different endings. Here, it is described how the results of these screens facilitated the development of novel compounds to be used in the process of isolating and sequencing of this non‐canonical mGluR. This study shows how the compounds were tested for their ability to alter stretch‐evoked afferent firing in muscle spindles and SLV endocytosis in the lanceolate endings of hair follicles to ensure they maintained their ability to bind to the receptor. For the development of novel compounds, kainate was chosen as the parent ligand due to its potency and ease of chemical modification. Novel kainate derivatives were then synthesised and tested to find potent analogues suitable for ‘click‐chemistry’, an established technique for relatively quick, cheap, stereospecific and high‐yield chemical modifications (Angewandte Chemie (International ed. in English), 40, 2001, pp2004). Of the novel kainate analogues developed, unfortunately ZCZ49 and ZCZ50 lost the ability to produce a significant change in spindle stretch‐evoked firing. However, ZCZ90 was as potent as kainate, increasing firing by a similar margin at 1 μm (n = 8; P < 0.001). The addition of either a biotin or a fluorescein side group to ZCZ90, using the click‐chemistry technique, did not affect the potency and hence these compounds will be used in further studies of the receptor. As well as the development of these compounds, the study found not only many similarities, but also some key differences between the two types of primary mechanosensory endings investigated. These differences must be taken into account in further study. However, they also present an intriguing opportunity for these receptors to be targeted selectively to modulate ending sensitivity as treatments for muscle spasm in multiple sclerosis and spinal cord injury, and possibly even baroreceptor firing to treat hypertension.     

Distinct trafficking and expression mechanisms underlie LTP and LTD of NMDA receptor‐mediated synaptic responses

Although an increasing number of studies have demonstrated the plasticity of NMDA receptor‐mediated synaptic transmission, little is known about the molecular mechanisms that underlie this neurologically important process. In a study of NMDAR‐mediated synaptic responses in hippocampal Schaffer‐CA1 synapses whose AMPA receptor (AMPAR) activity is totally blocked, we uncovered differences between the trafficking mechanisms that underlie the long‐term potentiation (LTP) and long‐term depression (LTD) that can be induced in these cells under these conditions. The LTP‐producing protocol failed to induce a change in the amplitude of NMDAR‐mediated postsynaptic currents (NMDAR EPSCs) in the first 5–10 min, but induced gradual enhancement of NMDAR EPSCs thereafter that soon reached a stable magnitude. This “slow” LTP of NMDAR EPSCs (LTP_NMDA) was blocked by inhibiting exocytosis or actin polymerization in postsynaptic cells. By contrast, LTD of NMDAR EPSCs (LTD_NMDA) was immediately inducible, and, although it was blocked by the actin stabilizer, it was unaffected by exocytosis or endocytosis inhibitors. Furthermore, concomitant changes in the decay time of NMDAR EPSCs suggested that differential switches in NR2 subunit composition accompanied LTP_NMDA and LTD_NMDA, and these changes were blocked by the calcium buffer BAPTA or an mGluR antagonist. Our results suggest that LTP_NMDA and LTD_NMDA utilize different NMDAR trafficking pathways and express different ratios of NMDAR subunits on the postsynaptic surface. © 2009 Wiley‐Liss, Inc.     

Elevated glycogen synthase kinase-3 activity in Fragile X mice: key metabolic regulator with evidence for treatment potential

Significant advances have been made in understanding the underlying defects of and developing potential treatments for Fragile X syndrome (FXS), the most common heritable mental retardation. It has been shown that neuronal metabotropic glutamate receptor 5 (mGluR5)-mediated signaling is affected in FX animal models, with consequent alterations in activity-dependent protein translation and synaptic spine functionality. We demonstrate here that a central metabolic regulatory enzyme, glycogen synthase kinase-3 (GSK3) is present in a form indicating elevated activity in several regions of the FX mouse brain. Furthermore, we show that selective GSK3 inhibitors, as well as lithium, are able to revert mutant phenotypes of the FX mouse. Lithium, in particular, remained effective with chronic administration, although its effects were reversible even when given from birth. The combination of an mGluR5 antagonist and GSK3 inhibitors was not additive. Instead, it was discovered that mGluR5 signaling and GSK3 activation in the FX mouse are coordinately elevated, with inhibition of mGluR5 leading to inhibition of GSK3. These findings raise the possibility that GSK3 is a fundamental and central component of FXS pathology, with a substantial treatment potential.     

Interactions between the mGluR2/3 agonist, LY379268, and cocaine on in vivo neurochemistry and behavior in squirrel monkeys

Recent evidence indicates that group II metabotropic glutamate receptors (mGluR2 and mGluR3) may play a role in the pathology of cocaine addiction. The purpose of the current study was to determine the effects of the mGluR2/3 agonist, LY379268, on cocaine-induced changes in DA neurochemistry in nonhuman primates. Furthermore, the current study aimed to determine if changes in DA neurochemistry would correlate with LY379268-induced changes in the behavioral effects of cocaine. In vivo microdialysis was conducted in conscious squirrel monkeys (n = 4) in order to monitor cocaine-induced changes in extracellular DA in the caudate nucleus. Separate groups of subjects were trained on a fixed-interval schedule of stimulus termination (n = 4) or a second-order schedule of cocaine self-administration (n = 5) to characterize the behavioral-stimulant and reinforcing effects, respectively. LY379268 significantly attenuated cocaine-induced increases in DA. LY379268 also significantly attenuated cocaine-induced behavioral-stimulant effects following a short pretreatment time, but not following a longer pretreatment time. Cocaine self-administration was significantly attenuated but only at an intermediate pretreatment dose of LY379268. Moreover, reinstatement of previously extinguished cocaine self-administration was not significantly attenuated by LY379268. Hence, drug interactions on neurochemistry did not correlate well with behavioral measures.     

Unique presynaptic and postsynaptic roles of Group II metabotropic glutamate receptors in the modulation of thalamic network activity

The thalamic reticular nucleus (TRN) is a sheet of GABAergic neurons that project to other TRN neurons and to associated thalamocortical relay nuclei. The TRN receives glutamatergic synaptic inputs from cortex as well as reciprocal inputs from the collaterals of thalamocortical neurons. In addition to ionotropic glutamate receptors, metabotropic glutamate receptors (mGluRs) are present in the TRN circuitry. Using whole cell voltage clamp recordings, we pharmacologically characterized unique pre- and postsynaptic functions for Group II mGluRs (mGluR 2 and mGluR 3) within the TRN circuitry in ferrets. mGluR 2 was found on presynaptic cortical axon terminals in the TRN, where it reduced glutamate release, while mGluR 3 acted postsynaptically on TRN cells to increase membrane conductance. Using miniature inhibitory postsynaptic current analysis, we also found that picrotoxin-sensitive intra-TRN GABA-mediated neurotransmission was not affected by administration of a Group II mGluR agonist, indicating that neither mGluR 2 nor 3 acts on presynaptic GABA-containing terminals within the TRN. Because strong corticothalamic activation is implicated in abnormal thalamic rhythms, we used extracellular recordings in the lateral geniculate nucleus to study the effect of Group II mGluR agonists upon these slow oscillations. We induced approximately 3 Hz spike-and-wave discharge activity through corticothalamic stimulation, and found that such activity was reduced in the presence of the Group II mGluR agonist, (-)-2-oxa-4-aminobicyclo[3.1.0]hexane-4,6-dicarboxylate (LY379268). These data indicate that Group II mGluR reduce the impact of corticothalamic excitation, and that they may be a useful target in the reduction of absence-like rhythms.     

Effect of the metabotropic glutamate receptor type 5 antagonists MPEP and MTEP in parkinsonian monkeys

Brain glutamate overactivity is well documented in Parkinson's disease (PD) and antiglutamatergic drugs have been proposed to relieve PD symptoms and decrease dyskinesias. Metabotropic glutamate receptors are topics of recent interest in PD. This study investigated the effects of the metabotropic glutamate receptors type 5 (mGluR5) antagonists MPEP and MTEP on motor behavior in monkeys with a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) lesion to model PD and treated with l-Dopa the gold standard therapy. Six Macaca fascicularis MPTP monkeys were initially treated repeatedly with l-Dopa; this treatment increased their locomotion and reduced their parkinsonian scores but also induced dyskinesias. Then, a dose-response of MPEP and MTEP (1.5-30 mg/kg) administered 15 and 30 min respectively prior to l-Dopa, showed that the antiparkinsonian activity of l-Dopa was generally maintained as measured with locomotion and antiparkinsonian scores as well as the onset and duration of the l-Dopa response. Interestingly the mean dyskinesia score during all the duration of the l-Dopa motor effect, the 1 h peak period dyskinesias scores as well as the maximal dyskinesias scores were dose-dependently reduced with both drugs reaching statistical significance at 10 and 30 mg/kg. Our results showed a beneficial antidyskinetic effect of blocking mGluR5 in l-Dopa-treated MPTP monkeys. This supports the therapeutic use of an mGluR5 antagonist to restore normal brain glutamate neurotransmission in PD and decrease dyskinesias.     

Glutamatergic targets for enhancing extinction learning in drug addiction

The persistence of the motivational salience of drug-related environmental cues and contexts is one of the most problematic obstacles to successful treatment of drug addiction. Behavioral approaches to extinguishing the salience of drug-associated cues, such as cue exposure therapy, have generally produced disappointing results which have been attributed to, among other things, the context specificity of extinction and inadequate consolidation of extinction learning. Extinction of any behavior or conditioned response is a process of new and active learning, and increasing evidence suggests that glutamatergic neurotransmission, a key component of the neural plasticity that underlies normal learning and memory, is also involved in extinction learning. This review will summarize findings from both animal and human studies that suggest that pharmacological enhancement of glutamatergic neurotransmission facilitates extinction learning in the context of drug addiction. Pharmacological agents that have shown potential efficacy include NMDA partial agonists, mGluR5 receptor positive allosteric modulators, inhibitors of the GlyT1 glycine transporter, AMPA receptor potentiators, and activators of the cystine-glutamate exchanger. These classes of cognition-enhancing compounds could potentially serve as novel pharmacological adjuncts to cue exposure therapy to increase success rates in attenuating cue-induced drug craving and relapse.     

Activation of group II metabotropic glutamate receptors reduces directional selectivity in retinal ganglion cells

In the mammalian retina, high levels of the group II metabotropic glutamate receptor (mGluR) subtype are expressed in starburst amacrine cells. A prominent role of starburst amacrine cells is the generation of directional selectivity in ON-OFF directionally selective retinal ganglion cells (DS RGCs). Extracellular microelectrodes were used to study the effects of activation of group II mGluRs on the responses of rabbit ON-OFF DS RGCs to a moving light stimulus. Directionally selective responses in these RGCs were substantially reduced by the selective group II mGluR agonist DCG-IV. DCG-IV brought out a response to movement in the null direction that was similar in magnitude and time course to the response to movement in the preferred direction. This effect of DCG-IV was reversed by the group II mGluR antagonist EGLU. Application of EGLU alone failed to alter directional selectivity in the RGCs but did reduce the response to movement in the preferred direction. To determine whether group II mGluRs modulate the release of the neurotransmitter acetylcholine from starburst amacrine cells, the effect of DCG-IV on ON-OFF DS RGCs was examined in the presence of ambenonium, an acetylcholinesterase inhibitor. When applied alone, ambenonium greatly prolonged the responses of ON-OFF DS RGCs to a light stimulus. This effect of ambenonium was completely abolished upon application of DCG-IV. Overall, the results suggest that postsynaptic group II mGluRs have the potential to influence directional selectivity in RGCs by inhibiting transmitter release from starburst amacrine cells.     

Neuroprogesterone: key to estrogen positive feedback?

In the cycling female rat, estradiol and progesterone induce reproductive behavior and the surge of luteinizing hormone (LH) needed for ovulation. Circulating estradiol of ovarian origin induces progesterone receptors in the preoptic area and hypothalamus. Sequential activation of estrogen receptors (ER) and progesterone receptors coordinates reproductive physiology and behavior. In ovariectomized and adrenalectomized (ovx/adx) rats, administration of estradiol alone is sufficient to initiate an LH surge, and central infusion of aminoglutethimide (AGT), a blocker of the P450 side chain cleavage enzyme, disrupted the estrous cycle of intact rats without affecting peripheral estradiol levels, suggesting that an endogenous source of progesterone remains in these animals. In ovx/adx rats, progesterone levels in the hypothalamus increase prior to the LH surge, and inhibition of progesterone synthesis prevents the LH surge, suggesting that hypothalamic neuroprogesterone is necessary for estrogen positive feedback. In support of the idea that estradiol induces neuroprogesterone, estradiol increased expression of the progesterone-synthesizing enzyme 3β-hydroxysteroid dehydrogenase (3β-HSD) in the hypothalamus before the LH surge. Further, in vitro experiments demonstrate that estradiol stimulates progesterone synthesis in astrocytes, considered to be the most active steroidogenic cells in the CNS. To stimulate neurosteroidogenesis, estradiol acts through membrane ER and type 1a metabotropic glutamate receptors (mGluR1a) to increase free cytoplasmic calcium ([Ca²⁺]_i) via activation of the PLC–IP₃ pathway. Estradiol-induced progesterone synthesis is mimicked by thapsigargin-induced release of IP₃ receptor-sensitive Ca²⁺ stores in astrocyte cultures. Thus, estradiol-induced progesterone synthesis is dependent on membrane ERs that act through mGluR1a to activate the PLC–IP₃ pathway. This neuroprogesterone also facilitated proceptive behavior. Blocking either progesterone synthesis or progesterone receptor in estrogen-primed ovx/adx prevented proceptive but not receptive behaviors.     

Endocannabinoid signaling in the spinal locomotor circuitry

To understand how the spinal central pattern generators produce locomotor movements, it is necessary to characterize the network's connectivity, the intrinsic properties of the constituent neurons and the modulatory mechanisms. Modulation operating within spinal locomotor networks is required for the generation of the final motor output. In this review, we have summarized how endocannabinoids released by locomotor network neurons contribute to setting the baseline locomotor frequency. They are synthesized on demand as a result of activation of mGluR1 and act as retrograde messengers to depress inhibitory synaptic transmission. We also discuss how endogenous activation of mGluR1 contributes to the normal operation of the spinal locomotor network and the underlying cellular and synaptic mechanisms.