Behavioral stress is recognized as a main risk factor for neuropsychiatric diseases. Converging evidence suggested that acute stress is associated with increase of excitatory transmission in certain forebrain areas. Aim of this work was to investigate the mechanism whereby acute stress increases glutamate release, and if therapeutic drugs prevent the effect of stress on glutamate release. Rats were chronically treated with vehicle or drugs employed for therapy of mood/anxiety disorders (fluoxetine, desipramine, venlafaxine, agomelatine) and then subjected to unpredictable footshock stress. Acute stress induced marked increase in depolarization-evoked release of glutamate from synaptosomes of prefrontal/frontal cortex in superfusion, and the chronic drug treatments prevented the increase of glutamate release. Stress induced rapid increase in the circulating levels of corticosterone in all rats (both vehicle- and drug-treated), and glutamate release increase was blocked by previous administration of selective antagonist of glucocorticoid receptor (RU 486). On the molecular level, stress induced accumulation of presynaptic SNARE complexes in synaptic membranes (both in vehicle- and drug-treated rats). Patch-clamp recordings of pyramidal neurons in the prefrontal cortex revealed that stress increased glutamatergic transmission through both pre- and postsynaptic mechanisms, and that antidepressants may normalize it by reducing release probability. Acute footshock stress up-regulated depolarization-evoked release of glutamate from synaptosomes of prefrontal/frontal cortex. Stress-induced increase of glutamate release was dependent on stimulation of glucocorticoid receptor by corticosterone. Because all drugs employed did not block either elevation of corticosterone or accumulation of SNARE complexes, the dampening action of the drugs on glutamate release must be downstream of these processes. This novel effect of antidepressants on the response to stress, shown here for the first time, could be related to the therapeutic action of these drugs.

Glutamate neurotransmission was recently implicated in the action of stress and in antidepressant mechanisms. We report that chronic (not acute) treatment with three antidepressants with different primary mechanisms (fluoxetine, reboxetine, and desipramine) markedly reduced depolarization-evoked release of glutamate, stimulated by 15 or 25 mmKCl, but not release of GABA. Endogenous glutamate and GABA release was measured in superfused synaptosomes, freshly prepared from hippocampus of drug-treated rats. Interestingly, treatment with the three drugs only barely changed the release of glutamate (and of GABA) induced by ionomycin. In synaptic membranes of chronically treated rats we found a marked reduction in the protein-protein interaction between syntaxin 1 and Thr²⁸⁶-phosphorylated αCaM kinase II (α-calcium/calmodulin-dependent protein kinase II) (an interaction previously proposed to promote neurotransmitter release) and a marked increase in the interaction between syntaxin 1 and Munc-18 (an interaction proposed to reduce neurotransmitter release). Furthermore, we found a selective reduction in the expression level of the three proteins forming the core SNARE (solubleN-ethylmaleimide-sensitive factor attachment protein receptor) complex. These findings suggest that antidepressants work by stabilizing glutamate neurotransmission in the hippocampus and that they may represent a useful tool for the study of relationship between functional and molecular processes in nerve terminals.

Despite some advances in the understanding of amyotrophic lateral sclerosis (ALS) pathogenesis, significant achievements in treating this disease are still lacking. Mesenchymal stromal (stem) cells (MSCs) have been shown to be effective in several models of neurological disease. To determine the effects of the intravenous injection of MSCs in an ALS mouse model during the symptomatic stage of disease, MSCs (1 × 10⁶) were intravenously injected in mice expressing human superoxide dismutase 1 (SOD1) carrying the G93A mutation (SOD1/G93A) presenting with experimental ALS. Survival, motor abilities, histology, oxidative stress markers and [³H]D-aspartate release in the spinal cord were investigated. MSC injection in SOD1/G93A mice improved survival and motor functions compared with saline-injected controls. Injected MSCs scantly home to the central nervous system and poorly engraft. We observed a reduced accumulation of ubiquitin agglomerates and of activated astrocytes and microglia in the spinal cord of MSC-treated SOD1/G93A mice, with no changes in the number of choline acetyltransferase- and glutamate transporter type 1-positive cells. MSC administration turned around the upregulation of metallothionein mRNA expression and of the activity of the antioxidant enzyme glutathione S-transferase, both associated with disease progression. Last, we observed that MSCs reverted both spontaneous and stimulus-evoked neuronal release of [³H]D-aspartate, a marker of endogenous glutamate, which is upregulated in SOD1/G93A mice. These findings suggest that intravenous administration of MSCs significantly improves the clinical outcome and pathological scores of mutant SOD1/G93A mice, thus providing the rationale for their exploitation for the treatment of ALS.

Alterations in the autophagic pathway are associated with the onset and progression of various diseases. However, despite the therapeutic potential for pharmacological modulators of autophagic flux, few such compounds have been characterised. Here we show that clomipramine, an FDA-approved drug long used for the treatment of psychiatric disorders, and its active metabolite desmethylclomipramine (DCMI) interfere with autophagic flux. Treating cells with DCMI caused a significant and specific increase in autophagosomal markers and a concomitant blockage of the degradation of autophagic cargo. This observation might be relevant in therapy in which malignant cells exploit autophagy to survive stress conditions, rendering them more susceptible to the action of cytotoxic agents. In accordance, DCMI-mediated obstruction of autophagic flux increased the cytotoxic effect of chemotherapeutic agents. Collectively, our studies describe a new function of DCMI that can be exploited for the treatment of pathological conditions in which manipulation of autophagic flux is thought to be beneficial.