Differential acetylcholine and GABA release from cultured chick retina cells.
In the present work we investigated the mechanisms controlling the release of acetylcholine (ACh) and of gamma-aminobutyric acid (GABA) from cultures of amacrine-like neurons, containing a subpopulation of cells which are simultaneously GABAergic and cholinergic. We found that 81.2 +/- 2.8% of the cells present in the culture were stained immunocytochemically with an antibody against choline acetyltransferase, and 38.5 +/- 4.8% of the cells were stained with an antibody against GABA. Most of the cells containing GABA (87.0 +/- 2.9%) were cholinergic. The release of acetylcholine and GABA was mostly Ca2+-dependent, although a significant release of [3H]GABA occurred by reversal of its transporter. Potassium evoked the Ca2+-dependent release of [3H]GABA and [3H]acetylcholine, with EC50 of 31.0 +/- 1.0 mm and 21.6 +/- 1.1 mm, respectively. The Ca2+-dependent release of [3H]acetylcholine was significantly inhibited by 1 micrometer tetrodotoxin and by low (30 nm) omega-conotoxin GVIA (omega-CgTx GVIA) concentrations, or by high (300 nm) nitrendipine (Nit) concentrations. On the contrary, the release of [14C]GABA was reduced by 30 nm nitrendipine, or by 500 nm omega-CgTx GVIA, but not by this toxin at 30 nm. The release of either transmitters was unaffected by 200 nm omega-Agatoxin IVA (omega-Aga IVA), a toxin that blocks P/Q-type voltage-sensitive Ca2+ channels (VSCC). The results show that Ca2+-influx through omega-CgTx GVIA-sensitive N-type VSCC and through Nit-sensitive L-type VSCC induce the release of ACh and GABA. However, the significant differences observed regarding the Ca2+ channels involved in the release of each neurotransmitter suggest that in amacrine-like neurons containing simultaneously GABA and acetylcholine the two neurotransmitters may be released in distinct regions of the cells, endowed with different populations of VSCC.
Santos
P F
PF
Center for Neuroscience of Coimbra, Department of Zoology, University of Coimbra, 3000 Coimbra, Portugal.
Carvalho
A L
AL
Carvalho
A P
AP
Duarte
C B
CB
D000109:Acetylcholine / Q000378:metabolism*; D000818:Animals; D002118:Calcium / Q000502:physiology; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D002642:Chick Embryo; D002799:Cholinergic Fibers / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D008564:Membrane Potentials / Q000502:physiology; D011189:Potassium Chloride / Q000494:pharmacology; D012160:Retina / Q000378:metabolism*; D014316:Tritium; D005680:gamma-Aminobutyric Acid / Q000378:metabolism*
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D014316:Tritium; D005680:gamma-Aminobutyric Acid; D011189:Potassium Chloride; D000109:Acetylcholine; D002118:Calcium