Evaluation of the Safety, Tolerability, and Pharmacokinetic Profiles of TP0473292 (TS-161), A Prodrug of a Novel Orthosteric mGlu2/3 Receptor Antagonist TP0178894, in Healthy Subjects and Its Antidepressant-Like Effects in Rodents.

BACKGROUND: TP0473292 (the active ingredient of TS-161) is a prodrug of a novel metabotropic glutamate (mGlu) 2/3 receptor antagonist being developed for the treatment of patients with depression. This study evaluated the safety, tolerability, and pharmacokinetics of orally administered TS-161 in healthy subjects. METHODS: This was a first-in-human, phase 1, randomized, double-blind, placebo-controlled, single-ascending dose (15-400 mg TS-161) and 10-day multiple-ascending dose (50-150 mg TS-161) study in healthy subjects, conducted from June 2019 through February 2020. Plasma and urine concentrations of the prodrug and its metabolites, and cerebrospinal fluid (CSF) concentrations of the active metabolite TP0178894 were measured to evaluate the pharmacokinetic profiles after oral administration of TS-161. RESULTS: Following single and multiple doses, TP0473292 was extensively converted into its active metabolite TP0178894. Plasma concentrations of TP0178894 reached peak (Cmax) within 5 hours post dose and declined with a t1/2 <13 hours. Plasma exposures of TP0178894 increased with increasing dose. TP0178894 penetrated into CSF and reached a Cmax of 9.892 ng/mL at a single dose of 100 mg, which was comparable with IC50 values of antagonist activity at mGlu2/3 receptors. The most frequently observed adverse events that showed exposure-related incidence during the study were nausea, vomiting, and dizziness. CONCLUSIONS: The mGlu2/3 receptor antagonist prodrug TP0473292 is safe and well-tolerated, is orally bioavailable in humans with extensive conversion into the active metabolite TP0178894 with sufficient CSF penetration to exert the anticipated pharmacological effects, and is a promising candidate for further clinical development in treatment of patients with depression.

Positive allosteric modulators (PAMs) of the group II metabotropic glutamate receptors: Design, synthesis, and evaluation as ex-vivo tool compounds.

This letter describes synthesis and evaluation of two series of dual mGlu2/mGlu3 positive allosteric modulators with moderate mGlu3 potency and robust mGlu2 potency in thallium flux assays. These compounds were profiled their ability to modulate mGlu3-mediated signaling in central neurons by co-application of a selective mGlu2 NAM to isolate mGlu3-selective effects. Using acute mouse brain slices from the prefrontal cortex, potentiation of group II mGlu receptor agonist Ca2+ signaling in PFC pyramidal cells with either the dual mGlu2/mGlu3 PAM 16e or 23d demonstrated effects mediated selectively via mGlu3.

Restoring the ON Switch in Blind Retinas: Opto-mGluR6, a Next-Generation, Cell-Tailored Optogenetic Tool.

Photoreceptor degeneration is one of the most prevalent causes of blindness. Despite photoreceptor loss, the inner retina and central visual pathways remain intact over an extended time period, which has led to creative optogenetic approaches to restore light sensitivity in the surviving inner retina. The major drawbacks of all optogenetic tools recently developed and tested in mouse models are their low light sensitivity and lack of physiological compatibility. Here we introduce a next-generation optogenetic tool, Opto-mGluR6, designed for retinal ON-bipolar cells, which overcomes these limitations. We show that Opto-mGluR6, a chimeric protein consisting of the intracellular domains of the ON-bipolar cell-specific metabotropic glutamate receptor mGluR6 and the light-sensing domains of melanopsin, reliably recovers vision at the retinal, cortical, and behavioral levels under moderate daylight illumination.

The effects of a novel metabotropic glutamate receptor 5 antagonist (AZD2066) on transient lower oesophageal sphincter relaxations and reflux episodes in healthy volunteers.

BACKGROUND: Selective metabotropic glutamate receptor 5 (mGluR5) antagonists inhibit transient lower oesophageal sphincter relaxations (TLESRs) in animals and acid reflux in humans. AIM: To assess the effect of single doses of the mGluR5 antagonist AZD2066 on TLESRs and reflux in humans. METHODS: Healthy male volunteers received AZD2066 13 mg and placebo (part A), or AZD2066 2 mg and AZD2066 6 mg and placebo (part B), in a randomised crossover study. Postprandial manometry/pH-impedance measurements were taken after each dose. RESULTS: A total of 13 individuals completed part A of the study and 19 individuals completed part B. There was a significant reduction in the geometric mean number of TLESRs (27%; P = 0.02) and the geometric mean number of reflux episodes (51%; P = 0.01) in subjects receiving AZD2066 13 mg compared with placebo. Adverse events in participants receiving AZD2066 13 mg were mostly related to the nervous system [dizziness (3/13); disturbance in attention (3/13)]. Adverse events were reversible and of mild intensity. There were no serious adverse events. The effects of AZD2066 appeared dose-dependent, with smaller reductions in TLESRs and reflux episodes (relative to placebo) and fewer adverse events observed for AZD2066 2 mg and AZD2066 6 mg compared with AZD2066 13 mg. CONCLUSION: The mGluR5-mediated inhibition of TLESRs may be a useful approach for inhibiting gastro-oesophageal reflux.

Randomised clinical trial: effects of monotherapy with ADX10059, a mGluR5 inhibitor, on symptoms and reflux events in patients with gastro-oesophageal reflux disease.

BACKGROUND: ADX10059, a metabotropic glutamate receptor 5 (mGluR5) negative allosteric modulator, has been shown to reduce gastro-oesophageal reflux events and oesophageal acid exposure in patients with gastro-oesophageal reflux disease (GERD) and healthy subjects. AIM: To evaluate the effects of ADX10059 monotherapy for 2 weeks on symptom control in patients with GERD. METHODS: This was a double-blind, placebo-controlled, multi-centre trial in GERD patients who were responders to proton pump inhibitors (PPIs). Following PPIs withdrawal, a 2-week baseline washout period was followed by 2-week treatment with either ADX10059 120 mg or placebo b.d. The primary clinical efficacy endpoint was the number of GERD symptom-free days in treatment week 2 compared with the last 7 days of baseline. The effect on reflux events using 24-h impedance-pH monitoring was also determined in a subset of 24 patients. RESULTS: The full analysis set comprised 103 patients ADX10059 (N= 50), Placebo (N=53). In treatment week 2, ADX10059 significantly increased GERD symptom-free days (P=0.045) and heartburn-free days (P=0.037), reduced antacid use (P=0.017), improved total symptom score (P=0.048) including subscale heartburn/regurgitation (P=0.007) and sleep disturbance because of GERD (P= 0.022). ADX10059 significantly reduced total (P=0.034) and acidic reflux events (P=0.003). ADX10059 was well tolerated. Most common adverse events for ADX10059 were mild to moderate dizziness 16% and vertigo 12% (placebo 4% and 2%). CONCLUSIONS: Inhibition of mGluR5 with ADX10059 monotherapy reduces reflux events and improves symptoms in GERD patients. This mechanism has promise for the management of GERD.

Subcellular and subsynaptic localization of group I metabotropic glutamate receptors in the nucleus accumbens of cocaine-treated rats.

There is significant pharmacological and behavioral evidence that group I metabotropic glutamate receptors (mGluR1a and mGluR5) in the nucleus accumbens play an important role in the neurochemical and pathophysiological mechanisms that underlie addiction to psychostimulants. To further address this issue, we undertook a detailed ultrastructural analysis to characterize changes in the subcellular and subsynaptic localization of mGluR1a and mGluR5 in the core and shell of nucleus accumbens following acute or chronic cocaine administration in rats. After a single cocaine injection (30 mg/kg) and 45 min withdrawal, there was a significant decrease in the proportion of plasma membrane-bound mGluR1a in accumbens shell dendrites. Similarly, the proportion of plasma membrane-bound mGluR1a was decreased in large dendrites of accumbens core neurons following chronic cocaine exposure (i.e. 1-week treatment followed by 3-week withdrawal). However, neither acute nor chronic cocaine treatments induced significant change in the localization of mGluR5 in accumbens core and shell, which is in contrast with the significant reduction of plasma membrane-bound mGluR1a and mGluR5 induced by local intra-accumbens administration of the group I mGluR agonist, (RS)-3,5-dihydroxyphenylglycine (DHPG). In conclusion, these findings demonstrate that cocaine-induced glutamate imbalance has modest effects on the trafficking of group I mGluRs in the nucleus accumbens. These results provide valuable information on the neuroadaptive mechanisms of accumbens group I mGluRs in response to cocaine administration.

Systemic and central amygdala injections of the mGluR(2/3) agonist LY379268 attenuate the expression of incubation of cocaine craving.

BACKGROUND: We and others reported time-dependent increases in cue-induced cocaine seeking after withdrawal, suggesting that craving incubates over time. Recently, we found that central amygdala extracellular signal-regulated kinases (ERK) and glutamate are involved in this incubation. Here, we further explored the role of central amygdala glutamate in the incubation of cocaine craving by determining the effect of systemic or central amygdala injections of the mGluR2/3 agonist LY379268 (which decreases glutamate release) on cue-induced cocaine seeking during early and late withdrawal. METHODS: Rats were trained to self-administer cocaine for 10 days (6 hours/day); infusions were paired with a tone-light cue. Cocaine seeking and craving after systemic or central amygdala injections of LY379268 were then assessed in extinction tests in the presence of the cocaine-associated cues during early (day 3) or late (day 21) withdrawal. RESULTS: Systemic (1.5 or 3 mg/kg) or central amygdala (.5 or 1.0 microg/side) injections of LY379268 attenuated enhanced extinction responding on day 21 but had no effect on lower extinction responding on day 3. CONCLUSIONS: Results confirm our previous findings on the role of central amygdala glutamate in the incubation of cocaine craving and together with previous reports suggest that mGluR(2/3) agonists should be considered in the treatment of drug relapse.

Metabotropic glutamate receptors and neuroadaptation to antidepressants: imipramine-induced down-regulation of beta-adrenergic receptors in mice treated with metabotropic glutamate 2/3 receptor ligands.

Antidepressant drugs have a clinical latency that correlates with the development of neuroadaptive changes, including down-regulation of beta-adrenergic receptors in different brain regions. The identification of drugs that shorten this latency will have a great impact on the treatment of major depressive disorders. We report that the time required for the antidepressant imipramine to reduce the expression of beta-adrenergic receptors in the hippocampus is reduced by a co-administration with centrally active ligands of type 2/3 metabotropic glutamate (mGlu2/3) receptors. Daily treatment of mice with imipramine alone (10 mg/kg, i.p.) reduced the expression of beta-adrenergic receptors in the hippocampus after 21 days, but not at shorter times, as assessed by western blot analysis of beta1-adrenergic receptors and by the amount of specifically bound [3H]CGP-12177, a selective beta-adrenergic receptor ligand. Down-regulation of beta-adrenergic receptors occurred at shorter times (i.e. after 14 days) when imipramine was combined with low doses (0.5 mg/kg, i.p.) of the selective mGlu2/3 receptor agonist LY379268, or with the preferential mGlu2/3 receptor antagonist LY341495 (1 mg/kg, i.p.). Higher doses of LY379268 (2 mg/kg, i.p.) were inactive. This intriguing finding suggests that neuroadaptation to imipramine--at least as assessed by changes in the expression of beta1-adrenergic receptors--is influenced by drugs that interact with mGlu2/3 receptors and stimulates further research aimed at establishing whether any of these drugs can shorten the clinical latency of classical antidepressants.

Chronic exposure to ammonia alters the modulation of phosphorylation of microtubule-associated protein 2 by metabotropic glutamate receptors 1 and 5 in cerebellar neurons in culture.

Hyperammonemia impairs signal transduction associated to glutamate receptors and phosphorylation of some neuronal proteins including microtubule-associated protein 2 (MAP-2). The aim of this work was to analyze the effects of hyperammonemia on modulation of MAP-2 phosphorylation by metabotropic glutamate receptors (mGluRs) in rat cerebellar neurons in culture. Hyperammonemia increased basal phosphorylation of MAP-2 (180%). Activation of mGluRs 1 and 5 with (S)-3,5-dihydroxyphenylglycine (DHPG) increased MAP-2 phosphorylation (170%) in control neurons but not in neurons exposed to ammonia. Activation of mGluRs 2 and 3 with (2S,3S,4S)-CCG/(2S, 1'S,2'S)-2-(carboxycyclopropyl)glycine increased slightly (25%) MAP-2 phosphorylation in neurons exposed to ammonia or not. Activation of mGluR5 with (+/-)-trans-azetidine-2,4-dicarboxylic acid increased MAP-2 phosphorylation (24%) in control neurons but decreased it by 56% in neurons exposed to ammonia. Activation of mGluR1 using 2-methyl-6-(phenylethynyl)pyridine and DHPG increased MAP-2 phosphorylation 183% in control neurons but only 89% in neurons exposed to ammonia. In control neurons mGluR1 activation greatly increases phosphorylation of MAP-2, while activation of mGluRs 5, 2 or 3 increased it slightly. Taken together, hyperammonemia reduces the increase in MAP-2 phosphorylation induced by mGluR1activation. Moreover, in neurons exposed to ammonia activation of mGluR5 reduces MAP-2 phosphorylation. These effects reflect significant alterations in signal transduction associated to mGluR1 and mGluR5 in hyperammonemia that may contribute to altered glutamatergic neurotransmission and to the neurological alterations in hyperammonemia and hepatic encephalopathy.

The anxiolytic-like activity of AIDA (1-aminoindan-1,5-dicarboxylic acid), an mGLu 1 receptor antagonist.

In the present study we examined the effects of 1-aminoindan-1,5-dicarboxylic acid (AIDA), regarded as a selective and competitive mGluR1 antagonist, in animal models of anxiety. Diazepam (1-10 mg/kg) was used as a reference drug. After intraperitoneal administration, AIDA (0.5-2 mg/kg) produced anxiolytic-like effects in the conflict drinking test and the elevated plus-maze test in rats; however, in doses up to 8 mg/kg, it was inactive in the four-plate test in mice. AIDA tested at the effective doses in the conflict drinking test changed neither the treshold current nor water intake in rats compared to vehicle treatment. AIDA (in a dose of 4 mg/kg, but not lower) increased the exploratory locomotor activity of rats measured in the open-field test, but it did not disturb rat motor coordination in the rota-rod test. The above results indicate that selective mGluR1 antagonist AIDA induces antianxiety-like effects at a low risk of acute side effects characteristic of benzodiazepines. Further studies are required to identify the sites and the mechanism of action of AIDA.

Activation of group III metabotropic glutamate receptors in selected regions of the basal ganglia alleviates akinesia in the reserpine-treated rat.

1. This study examined whether group III metabotropic glutamate (mGlu) receptor agonists injected into the globus pallidus (GP), substantia nigra pars reticulata (SNr) or intracerebroventricularly (i.c.v.) could reverse reserpine-induced akinesia in the rat. 2. Male Sprague-Dawley rats, cannulated above the GP, SNr or third ventricle, were rendered akinetic with reserpine (5 mg kg(-1) s.c.). 18 h later, behavioural effects of the group III mGlu receptor agonists L-serine-O-phosphate (L-SOP) or L-(+)-2-amino-4-phosphonobutyric acid (L-AP4) were examined. 3. In reserpine-treated rats, unilateral injection of L-SOP (2000 and 2500 nmol in 2.5 microl) into the GP produced a significant increase in net contraversive rotations compared to vehicle, reaching a maximum of 83+/-21 rotations 120 min(-1) (n=8). Pretreatment with the group III mGlu receptor antagonist methyl-serine-O-phosphate (M-SOP; 250 nmol in 2.5 microl) inhibited the response to L-SOP (2000 nmol) by 77%. Unilateral injection of L-SOP (250-1000 nmol in 2.5 microl) into the SNr of reserpine-treated rats produced a dose-dependent increase in net contraversive rotations, reaching a maximum of 47+/-6 rotations 30 min(-1) (n=6). M-SOP (50 nmol in 2.5 microl) inhibited the response to L-SOP (500 nmol) by 78%. 4. Following i.c.v. injection, L-SOP (2000-2500 nmol in 2.5 microl) or L-AP4 (0.5-100 nmol in 2 microl) produced a dose-dependent reversal of akinesia, attaining a maximum of 45+/-17 (n=8) and 72+/-3 (n=9) arbitrary locomotor units 30 min(-1), respectively. 6. These studies indicate that injection of group III mGlu receptor agonists into the GP, SNr or cerebral ventricles reverses reserpine-induced akinesia, the mechanism for which remains to be established.

Retrograde modulation of transmitter release by postsynaptic subtype 1 metabotropic glutamate receptors in the rat cerebellum.

1. The aim of the study was to elucidate the mechanisms underlying the depressant effect of the group I/II metabotropic glutamate receptor (mGluR) agonist 1S,3R-aminocyclopentane-1,3-dicarboxylic acid (1S,3R-ACPD) on parallel fibre (PF) to Purkinje cell (PC) synaptic transmission. Experiments were performed in rat cerebellar slices using the whole-cell patch-clamp technique and fluorometric measurements of presynaptic calcium variation 2. Analysis of short-term plasticity, fluctuation of EPSC amplitude and responses of PCs to exogenous glutamate showed that depression caused by 1S,3R-ACPD is presynaptic. 3. The effects of 1S,3R-ACPD were blocked and reproduced by group I mGluR antagonists and agonists, respectively. 4. These effects remained unchanged in mGluR5 knock-out mice and disappeared in mGluR1 knock-out mice. 5. 1S,3R-ACPD increased calcium concentration in PFs. This effect was abolished by AMPA/kainate (but not NMDA) receptor antagonists and mimicked by focally applied agonists of these receptors. Thus, it is not directly due to mGluRs but to presynaptic AMPA/kainate receptors indirectly activated by 1S,3R-ACPD. 6. Frequencies of spontaneous and evoked unitary EPSCs recorded in PCs were respectively increased and decreased by mGluR1 agonists. Similar results were obtained when mGluR1s were activated by tetanic stimulation of PFs. 7. Injecting 30 mM BAPTA into PCs blocked the effects of 1S,3R-ACPD on unitary EPSCs. 8. In conclusion, 1S,3R-ACPD reduces evoked release of glutamate from PFs. This effect is triggered by postsynaptic mGluR1s and thus implies that a retrograde messenger, probably glutamate, opens presynaptic AMPA/kainate receptors and consequently increases spontaneous release of glutamate from PF terminals and decreases evoked synaptic transmission.