Hippocampal astrocytes in situ respond to both exogenously-applied neurotransmitters and neuronally-released neurotransmitters.

摘要

A long standing question in neurobiology is whether or not astrocytes respond to the neuronal release of neurotransmitters in vivo. To address this question, acutely isolated hippocampal slices were loaded with calcium-sensitive dyes, and the responses of the astrocytes to exogenously-applied neuroligands and to electrical stimulation of the Schaffer collaterals were monitored by confocal microscopy. To confirm that the responsive cells were astrocytes the slices were immunostained for the astrocytic marker, glial fibrillary acidic protein.; The hippocampal astrocytes in situ responded to glutamate, kainate, {dollar}alpha{dollar}-amino-3-hydroxy-5-methylisoxazole-4-propionic acid, 1-aminocyclopentane-trans-1,3-dicarboxylic acid, N-methyl-D-aspartate, and depolarization with increases in the concentration of intracellular calcium ( (Ca{dollar}rmsp{lcub}2+{rcub}rbracksb{lcub}i{rcub}).{dollar} ATP and adenosine also induced dynamic increases in astrocytic (Ca{dollar}rmsp{lcub}2+{rcub}rbracksb{lcub}i{rcub}{dollar} which were blocked by the adenosine receptor antagonist, 8-(p-sulfophenyl)theophylline. The responses to adenosine were not blocked by tetrodotoxin, 8-cyclopentyltheophylline, 8-(3-chlorostyryl)caffeine, dipyridamole, or removal of extracellular calcium. The P{dollar}rmsb{lcub}2Y{rcub}{dollar} selective agonist, 2-methylthio-ATP, was unable to induce increases in astrocytic (Ca{dollar}rmsp{lcub}2+{rcub}rbracksb{lcub}i{rcub}{dollar} while the P{dollar}sb2{dollar} agonist {dollar}beta{dollar}-thio-ADP induced astrocytic responses in a very low percentage of astrocytes. These results indicate that hippocampal astrocytes in situ contain ionotropic and metabotropic glutamate receptors and P{dollar}sb1{dollar} purinergic receptors coupled to increases in (Ca{dollar}rmsp{lcub}2+{rcub}rbracksb{lcub}i{rcub}{dollar} while a small minority appear to contain P{dollar}sb2{dollar} purinergic receptors.; Stimulation of the Schaffer collaterals resulted in increases in (Ca{dollar}rmsp{lcub}2+{rcub}rbracksb{lcub}i{rcub}{dollar} in the astrocytes located in the stratum radiatum of CA1. The astrocytic responses were blocked by tetrodotoxin, {dollar}omega{dollar}-conotoxin MVIIC, and the selective metabotropic glutamate receptor antagonist, {dollar}alpha{dollar}-methyl-4-carboxyphenylglycine (MCPG). These results suggest that the astrocytic responses were induced by stimulation of metabotropic glutamate receptors on the astrocytes by neuronally released glutamate. The astrocytic responses to neuronal stimulation were enhanced in the presence of the K{dollar}sp+{dollar} channel antagonist, 4-aminopyridine (4-AP). Inhibition of the astrocytic responses in the presence of 4-AP required the presence of both MCPG and the ionotropic glutamate receptor antagonist, kynurenic acid. These results suggest that higher levels of neuronal activity result in stimulation of both metabotropic and ionotropic glutamate receptors on the astrocytes. Overall, the results indicate that hippocampal astrocytes in situ are able to respond to the neuronal release of the neurotransmitter glutamate with increases in (Ca{dollar}rmsp{lcub}2+{rcub}lbracksb{lcub}i{rcub}.{dollar}