Chronic stress enhances calcium mobilization and glutamate exocytosis in cerebrocortical synaptosomes from mice

Abstract

Objectives: Our previous study showed that acute restraint stress enhances depolarization-induced increases in intrasynaptosomal free calcium (Ca²⁺) concentration ([Ca²⁺]_i) and Ca²⁺-dependent glutamate release in mouse cerebrocortical nerve terminals (synaptosomes). In the present study, we investigated the effects of chronic stress on [Ca²⁺]_i and glutamate release in cerebrocortical synaptosomes from mice.

Methods: Male ddY strain mice were randomly assigned to one of two experimental groups: control group and chronic stressed group. Mice in the chronic stressed group were subjected to immobilization stress for 2 hours daily for a period of 21 days. [Ca²⁺]_i and glutamate release in cerebrocortical synaptosomes isolated from the mice were determined by fura-2 fluorescence assay and enzyme-linked fluorometric assay, respectively.

Results: Chronic stress caused a significant increase in resting [Ca²⁺]_i and significantly enhanced the ability of the depolarizing agents K⁺ and 4-aminopyridine (4-AP) to increase [Ca²⁺]_i. It also brought about a significant increase in spontaneous (unstimulated) glutamate release and significantly enhanced K⁺- and 4-AP-evoked Ca²⁺-dependent glutamate release. Synaptosomes were more sensitive to the depolarizing agents at lower concentrations following chronic stress than after acute stress. The pretreatment of synaptosomes with a combination of omega-agatoxin IVA (a P-type Ca²⁺ channel blocker) and omega-conotoxin GVIA (an N-type Ca²⁺ channel blocker) completely suppressed the enhancements of [Ca²⁺]_i and Ca²⁺-dependent glutamate release in chronic stressed mice.

Discussion: These results indicate that chronic stress enhances depolarization-evoked glutamate release by increasing [Ca²⁺]_i via stimulation of Ca²⁺ entry through P- and N-type Ca²⁺ channels, and that chronic stress increases the sensitivity to depolarizing agents.

Keywords:

• Ca2+ channel
• Cerebrocortical synaptosome
• Chronic stress
• Intrasynaptosomal free Ca2+
• Glutamate release