Monika Woźniak

Antidepressant-like effect of the mGluR5 antagonist MTEP in an astroglial degeneration model of depression

The glutamatergic predominance in the excitatory-inhibitory balance is postulated to be involved in the pathogenesis of depression. Such imbalance may be induced by astrocyte ablation which reduces glutamate uptake and increases glutamate level in the synaptic cleft. In the present study, we tried to ascertain whether astroglial degeneration in the prefrontal cortex could serve as an animal model of depression and whether inhibition of glutamatergic transmission by the mGluR5 antagonist MTEP could have antidepressant potential. Astrocytic toxins l-or dl-alpha-aminoadipic acid (AAA), 100μg/2μl, were microinjected, bilaterally into the rat medial prefrontal cortex (PFC) on the first and second day of experiment. MTEP (10mg/kg) or imipramine (30mg/kg) were administered on the fifth day. Following administration of MTEP or imipramine the forced swim test (FST) was performed for assessment of depressive-like behavior. The brains were taken out for analysis on day eight. The astrocytic marker, glial fibrillary acidic protein (GFAP) was quantified in PFC by Western blot method and by stereological counting of immunohistochemically stained sections. Both l-AAA and dl-AAA induced a significant increase in immobility time in the FST. This effect was reversed by imipramine, which indicates depressive-like effects of these toxins. A significant decrease in GFAP (about 50%) was found after l-AAA. Both the behavioral and GFAP level changes were prevented by MTEP injection. The obtained results indicate that the degeneration of astrocytes in the PFC by l-AAA may be a useful animal model of depression and suggest antidepressant potential of MTEP.

Glial degeneration as a model of depression

Major depression (MD) is a common and disabling disorder but knowledge of its pathophysiology is still incomplete. In the last years, degenerations or dysfunctions of glial cells, especially astrocytes, have been postulated to play a critical role in the pathogenesis of depression. Glial loss in prefrontal and limbic brain regions was observed in depressed patients and in animal models of stress and depression. Degeneration of astrocytes resulted in an excess glutamate in the synaptic cleft and glutamate/GABA imbalance in the affected structures. This review presents an up-to-date information concerning the role of glial cells in maintenance of glutamate/GABA balance in the brain tripartite glutamatergic synapses; discusses the importance of glial pathology and presents models of depression based on astrocyte impairment. The model of degeneration of astrocytes in the medial prefrontal cortex of the rat, induced by the specific astrocytic toxin α-aminoadipic acid, is presented as a valuable model for studying antidepressant compounds.

mGlu5-GABAB interplay in animal models of positive, negative and cognitive symptoms of schizophrenia

Diverse preclinical studies exploiting the modulation of the GABAergic and/or glutamatergic system in brain via metabotropic receptors suggest their potential therapeutic utility. GS39783 and CDPPB, positive allosteric modulators of GABAB and mGlu5 receptors, were previously shown to reverse behavioral phenotypes in animal models to mimic selected (predominantly positive) symptoms of schizophrenia. In the present study we investigated the activity of selected GABAB (GS39783 and CGP7930) and mGlu5 (CDPPB) positive allosteric modulators. We focused mainly on the aspects of their efficacy in the models of negative and cognitive symptoms of schizophrenia. We used modified swim test, social interactions (models of negative symptoms) and novel object recognition (model of cognitive disturbances). The activity of the compounds was also tested in haloperidol-induced catalepsy test. The mutual interaction between GABAB/mGlu5 ligands was investigated as well. In the second part of the study, DHPG-induced PI hydrolysis in the presence of GABAB receptor antagonist (SKF97541), and SKF97541-induced inhibition of cAMP formation in the presence of DHPG, was performed. Both mGlu5 and GABAB receptor modulators effectively reversed MK-801-induced deficits in behavioral models of schizophrenia. Moreover, the concomitant administration of sub-effective doses of CDPPB and GS39783 induced a clear antipsychotic-like effect in all the procedures used, except DOI-induced head twitches. The concomitant administration of group I mGlu and GABAB agonists did not displayed any synergistic effects in vitro. Summing up, an activation of both types of receptor may be a promising mechanism for the development of novel antipsychotic drugs, efficacious toward positive, negative and cognitive symptoms.

Involvement of GABA B Receptor Signaling in Antipsychotic-like Action of the Novel Orthosteric Agonist of the mGlu 4 Receptor, LSP4-2022

Considering that ligands of metabotropic glutamate and GABA receptors may exert beneficial effects on schizophrenia, we assessed the actions of the first mGlu4-selective orthosteric agonist, LSP4-2022, in several tests reflecting positive, negative, and cognitive symptoms of schizophrenia. Moreover, we investigated the possible involvement of GABAB receptors in LSP4-2022-induced actions. Hyperactivity induced by MK-801 or amphetamine and DOI-induced head twitches in mice were used as the models of positive symptoms. The social interaction test, modified forced swim test (FST), and novel object recognition (NOR) test were used as the models of negative and cognitive symptoms of schizophrenia. LSP4-2022 inhibited hyperactivity (in a dose-dependent manner, 0.5-2 mg/kg) induced by MK-801 or amphetamine and DOI-induced head twitches. In mGlu4 receptor knockout mice, LSP4-2022 was not effective. However, it reversed MK-801-induced impairment in the social interaction test and the MK-801-induced increase of immobility in the modified FST. In the NOR test, LSP4-2022 was active at a dose of 2 mg/kg. GABAB receptor antagonist, CGP55845 (10 mg/kg), reversed LSP4-2022-induced effects in hyperactivity and head twitch tests. At the same time, the simultaneous administration of subeffective doses of LSP4-2022 (0.1 mg/kg) and a positive allosteric modulator of GABAB receptor PAM, GS39783 (0.1 mg/kg), induced clear antipsychotic-like effects in those two tests. Such an interaction between mGlu4 and GABAB receptors was not observed in the social interaction and NOR tests. Therefore, we suggest that the activation of the mGlu4 receptor is a promising approach facilitating the discovery of novel antipsychotic drugs, and that the interplay between mGlu4 and GABAB receptors may become the basis for a novel therapy for schizophrenic patients with predomination of positive symptoms.

Neurochemical and behavioral studies on the 5-HT1A-dependent antipsychotic action of the mGlu4 receptor agonist LSP4-2022.

&NA; LSP4‐2022 is a novel, orthosteric agonist of mGlu4 receptor that induces antipsychotic‐like activity in animal studies. In the present study, the involvement of 5‐HT1A receptors in LSP4‐2022‐induced antipsychotic actions and the neurochemical background of that interaction were investigated. In several behavioral tests the actions of effective doses of the compound (0.5–2 mg/kg) were antagonized via the administration of the 5‐HT1A antagonist WAY100635 (0.1 mg/kg). The co‐administration of sub‐effective dose of the 5‐HT1A agonist (R)‐(S)‐8‐OH‐DPAT (0.01 mg/kg) intensified the activity of ineffective doses of LSP4‐2022, having no influence on the efficacy of the active doses. The co‐administration of effective doses of both compounds did not intensify each others action. In the microdialysis in vivo tests, MK‐801 (0.6 mg/kg) induced an enhancement of the release of dopamine, serotonin, glutamate and GABA in the prefrontal cortex. Administration of LSP4‐2022 (2 mg/kg) abolished this MK‐801‐induced effect on neurotransmitter release. Co‐administration with WAY100635 (0.1 mg/kg), a 5‐HT1A antagonist, completely (dopamine, serotonin) or partially (glutamate, GABA) counteracted this LSP4‐2022‐induced effect. Subsequently, the patch‐clamp recordings of spontaneous EPSCs were performed. sEPSCs were evoked in slices from the mouse prefrontal cortex by DOI (10 &mgr;M). LSP4‐2022 (2.5; 5 and 10 &mgr;m) reversed DOI‐induced changes in both the frequency and amplitude of the sEPSCs, but the more robust effect on the frequency was observed. The administration of WAY100635 had no effect on the LSP4‐2022‐induced effects on sEPSCs, indicating that the mGlu4‐5‐HT1A interaction does not occur via single‐neuron signaling but involves neuronal circuits that regulate neurotransmitter release. This article is part of the Special Issue entitled ‘Metabotropic Glutamate Receptors, 5 years on’. HighlightsLSP4‐2022 antagonizes MK‐801‐induced release of dopamine, serotonin, glutamate and GABAThe action of LSP4‐2022 is 5‐HT1A dependentmGlu4‐5‐HT1A interaction does not occur within a single neuron

Low-basicity 5-HT 7 Receptor Agonists Synthesized Using the van Leusen Multicomponent Protocol

A series of 5-aryl-1-alkylimidazole derivatives was synthesized using the van Leusen multicomponent reaction. The chemotype is the first example of low-basicity scaffolds exhibiting high affinity for 5-HT7 receptor together with agonist function. The chosen lead compounds 3-(1-ethyl-1H-imidazol-5-yl)-5-iodo-1H-indole (AGH-107, 1o, Ki 5-HT7 = 6 nM, EC50 = 19 nM, 176-fold selectivity over 5-HT1AR) and 1e (5-methoxy analogue, Ki 5-HT7 = 30 nM, EC50 = 60 nM) exhibited high selectivity over related CNS targets, high metabolic stability and low toxicity in HEK-293 and HepG2 cell cultures. A rapid absorption to the blood, high blood-brain barrier permeation and a very high peak concentration in the brain (Cmax = 2723 ng/g) were found for 1o after i.p. (5 mg/kg) administration in mice. The compound was found active in novel object recognition test in mice, at 0.5, 1 and 5 mg/kg. Docking to 5-HT7R homology models indicated a plausible binding mode which explain the unusually high selectivity over the related CNS targets. Halogen bond formation between the most potent derivatives and the receptor is consistent with both the docking results and SAR. 5-Chlorine, bromine and iodine substitution resulted in a 13, 27 and 89-fold increase in binding affinities, respectively, and in enhanced 5-HT1AR selectivity.

Mutual activation of glutamatergic mGlu4 and muscarinic M4 receptors reverses schizophrenia-related changes in rodents

RationaleMetabotropic glutamate receptors and muscarinic M4 receptors have been proposed as novel targets for various brain disorders, including schizophrenia. Both receptors are coupled to Go/i proteins and are expressed in brain circuits that are important in schizophrenia. Therefore, their mutual activation may be an effective treatment and allow minimizing the doses of ligands required for optimal activity.ObjectivesIn the present studies, subactive doses of mGlu4 and M4 activators (LSP4-2022 and VU152100, respectively) were administered to investigate the mutual interaction between mGlu4 and M4 receptors in animal models of schizophrenia.MethodsThe behavioral tests used were MK-801-induced hyperactivity, (±)-2.5-dimethoxy-4-iodoamphetamine hydrochloride (DOI)-induced head twitches, the modified forced swim test, and MK-801-induced disruptions of social interactions and novel object recognition. DOI-induced spontaneous excitatory postsynaptic currents (sEPSCs) in brain slices and positron emission tomography (PET) in were used to establish the ability of these compounds to modulate the glutamatergic and dopaminergic systems. Rotarod was used to assess putative adverse effects.ResultsThe mutual administration of subactive doses of LSP4-2022 and VU152100 exerted similar antipsychotic-like efficacy in animals as observed for active doses of both compounds, indicating their additive actions. VU152100 inhibited the DOI-induced frequency (but not amplitude) of sEPSCs in the frontal cortex, confirming presynaptic regulation of glutamate release. Both compounds reversed amphetamine-induced decrease in D2 receptor levels in the striatum, as measured with [18F]fallypride. The compounds did not induce any motor impartments when measured in rotarod test.ConclusionsBased on our results, the simultaneous activation of M4 and mGlu4 receptors is beneficial in reversing MK-801- and amphetamine-induced schizophrenia-related changes in animals.

Neurochemical changes underlying schizophrenia-related behavior in a modified forced swim test in mice

ABSTRACT The modified forced swim test (MFST) has excellent predictive validity for investigating the antipsychotic activity of drugs, with particular emphasis on their activity toward negative symptoms of schizophrenia. However, its face and construct validity are less understood. Therefore, in the present study, some biochemical changes within GABAergic and serotonergic neurotransmission that could be related to observed MK‐801‐induced disturbances and the activity of compounds active at those neurotransmitters were investigated. In biochemical experiments, mice were treated acutely or chronically with MK‐801 (13 days, 0.4 mg/kg). Their brains were dissected and frontal cortices and hippocampi were taken for further analysis. The levels of neurotransmitters were investigated with HPLC, and the expression of surrogate markers of schizophrenia (5‐HT1A receptors, GAD65, and GAD67, at both protein and mRNA levels) was measured via western blotting and qRT‐PCR. The modified forced swim test and locomotor activity were used to assess the activity of GABAB and 5‐HT1A‐related compounds. Repeated MK‐801 treatment (13 days, 0.4 mg/kg dose) led to decreases in the DOPAC/DA, 3MT/DA and HVA/DA metabolic ratios. Increased 5‐HT1A protein expression and decreased GAD65 and GAD67 protein expression was observed in both the cortex and hippocampus. mRNA levels for all proteins were decreased. The increased immobility in the forced swim test was reversed both by a GABAB agonist (SKF97541, 0.025 or 0.05 mg/kg), a positive allosteric modulator of GABAB receptor (racBHFF, 5 or 10 mg/kg) and by a 5‐HT1A agonist ((R)‐(+)‐8‐OH‐DPAT 0.01 or 0.025 mg/kg). Our research supports the hypothesis that changes in the levels of GABA and/or 5‐HT1A receptors may contribute to the schizophrenia‐like phenotype, and GABAergic and serotonergic agents may be good candidates for treating negative symptoms of schizophrenia. Highlights5‐HT1A receptor as a target for the treatment of negative symptoms of schizophreniaGABAB receptor as a target for the treatment of negative symptoms of schizophreniaChronic MK‐801 injections change the expression of surrogate markers of schizophrenia.

Negative Allosteric Modulators of mGlu7 Receptor as Putative Antipsychotic Drugs

The data concerning antipsychotic-like activity of negative allosteric modulators (NAMs)/antagonists of mGlu7 receptors are limited. The only available ligands for this receptor are MMPIP and ADX71743. In the present studies, we used stable cell line expressing mGlu7 receptor and it was shown that both compounds dose-dependently potentiated forskolin elevated cAMP concentration in the T-REx 293 cells, showing their inverse agonist properties. Subsequently, pharmacokinetic studies were performed. Both compounds were given intraperitoneally (i.p.) at the dose of 10 mg/kg and reached Cmax 0.25–0.5 h after administration, and then they declined rapidly, ADX71743 being almost undetectable 2 h after administration, while the concentration of MMPIP was still observed, suggesting that the concentration of MMPIP was more stable. Finally, we investigated the role of both mGlu7 receptor NAMs in animal models of schizophrenia. Behavioral tests commonly used in antipsychotic drug discovery were conducted. Both tested compounds dose-dependently inhibited MK-801-induced hyperactivity (MMPIP at 15 mg/kg; ADX at 5 and 15 mg/kg) and DOI-induced head twitches (MMPIP at 5, 10, 15 mg/kg; ADX at 2.5, 5, 10 mg/kg). Moreover, the same effects were noticed in novel object recognition test, where MMPIP (5, 10, 15 mg/kg) and ADX71743 (1, 5, 15 mg/kg) reversed MK-801-induced disturbances. In the social interaction test, antipsychotic activity was observed only for ADX71743 (5, 15 mg/kg). ADX71743 at the dose 2.5 mg/kg reversed MK-801-induced disruption in prepulse inhibition while MMPIP at 10 mg/kg reversed MK-801-induced disruption in spatial delayed alternation. The present studies showed that mGlu7 receptor may be considered as a putative target for antipsychotic drugs, though more studies are needed due to limited number of available ligands.

Simultaneous activation of muscarinic and GABAB receptors as a bidirectional target for novel antipsychotics

Abstract Recent preclinical studies point to muscarinic and GABAB receptors as novel therapeutic targets for the treatment of schizophrenia. This study was aimed to assess the role of muscarinic and GABAB receptor interactions in animal models of schizophrenia, using positive allosteric modulators (PAMs) of GABAB receptor (GS39783), muscarinic M4 (VU0152100) and M5 (VU0238429) receptor, and partial allosteric agonist of M1 receptor (VU0357017). DOI‐induced head twitches, social interaction and novel object recognition tests were used as the models of schizophrenia. Analyses of DOI‐induced increases in sEPSCs (spontaneous excitatory postsynaptic currents) were performed as complementary experiments to the DOI‐induced head twitch studies. Haloperidol‐induced catalepsy and the rotarod test were used to examine the adverse effects of the drugs. All three activators of muscarinic receptors were active in DOI‐induced head twitches. When administered together with GS39783 in subeffective doses, only the co‐administration of VU0152100 and GS39783 was effective. The combination also reduced the frequency but not the amplitude of DOI‐induced sEPSCs. Neither VU0357017 nor VU0238429 were active in social interaction test when given alone, and also the combination of VU0152100 and GS39783 failed to reverse MK‐801‐induced deficits observed in this test. All muscarinic activators when administered alone or in combination with GS39783 reversed the MK‐801‐induced disruption of memory in the novel object recognition test, and their actions were blocked by specific antagonists. None of the tested compounds or their combinations influenced the motor coordination of the animals. The compounds had no effect on haloperidol‐induced catalepsy and did not induce catalepsy when administered alone. Pharmacokinetic analysis confirmed lack of possible drug‐drug interactions after combined administration of GS39783 with VU0357017 or VU0152100; however, when the drug was co‐administered with VU0238429 its ability to pass the blood‐brain barrier slightly decreased, suggesting potential drug‐drug interactions. Our data show that modulation of cholinergic and GABAergic systems can potentially be beneficial in the treatment of the positive and cognitive symptoms of schizophrenia without inducing the adverse effects typical for presently used antipsychotics.