Evidence for two mechanisms of amino acid osmolyte release from hippocampal slices

A 30% decrease in osmolarity stimulated H-3-taurine, H-3-GABA and glutamate (followed as H-3-D-aspartate) efflux from rat hippocampal Slices. H-3-taur ine efflux was activated rapidly but inactivated slowly. It was decreased m arkedly by 100 muM 5 -nitro-(3-phenylpropylamino)benzoic acid (NPPB) and 60 0 muM niflumic acid and inhibited strongly by tyrphostins AG18, AG879 and A G 112 (25-100 muM), suggesting a tyrosine kinase-mediated mechanism. Hyposm olarity activated the mitogen-activated protein kinases (MAPK) extracellula r-signal-related kinase-1/2 (ERK1/ERK2) and p38, but blockade of this react ion did not affect H-3-taurine efflux. Hyposmosis also activated phosphatid ylinositide 3-kinase (PI3K) and its prevention by wortmannin (100 nM) essen tially abolished H-3-taurine efflUX. H-3-taurine efflux was insensitive to the protein kinase C (PKC) blocker chelerythrine (2.5 muM) or to cytochalas in E (3 muM). The release of H-3-GABA and H-3-D-aspartate occurred by a dif ferent mechanism, characterized by rapid activation and inactivation, insen sitivity to NPPB, niflumic acid, tyrphostins or wortmannin. H-3-GABA and H- 3-D-aspartate efflux was not due to external [NaCl] decrease, cytosolic Ca2 + increase or depolarization, or to reverse operation of the carrier. This novel mechanism of amino acid release may be mediated by Ca2+-independent e xocytosis and modulated by PKC and actin cytoskeleton disruption, as sugges ted by its inhibition by chelerythrine and potentiation by 100 nM phorbol-1 2-rayristate-13 acetate (PMA) and cytochalasin E. GABA and glutamate osmo-s ensitive efflux may explain the hyposmolarity-elicited increase in amplitud e of inhibitory and excitatory postsynaptic potentials in hippocampal slice s as well as the hyperexcitability associated with hyponatraemia.