New insights into molecular mechanism(s) underlying the presynaptic action of nitric oxide on GABA release.
BACKGROUND
Nitric oxide (NO) is an important presynaptic modulator of synaptic transmission. Here, we aimed to correlate the release of the major inhibitory neurotransmitter GABA with intracellular events occurring in rat brain axon terminals during their exposure to NO in the range of nanomolar-low micromolar concentrations.


METHODS
Using [(3)H]GABA and fluorescent dyes (Fluo 4-AM, acridine orange and rhodamine 6G), the following parameters were evaluated: vesicular and cytosolic GABA pools, intracellular calcium concentration, synaptic vesicle acidification, and mitochondrial membrane potential. Diethylamine NONOate (DEA/NO) and S-nitroso-N-acetylpenicillamine (SNAP) were used as NO donors.


RESULTS
DEA/NO and SNAP (in the presence of dithiothreitol (DTT)) stimulated external Ca(2+)-independent [(3)H]GABA release, which was not attributed to a rise in intracellular calcium concentration. [(3)H]GABA release coincided with increasing GABA level in cytosol and decreasing the vesicular GABA content available for exocytotic release. There was a strong temporal correlation between NO-induced increase in cytosolic [GABA] and dissipation of both synaptic vesicle proton gradient and mitochondrial membrane potential. Dissipation was reversible, and recovery of both parameters correlated in time with re-accumulation of [(3)H]GABA into synaptic vesicles. The molar ratio of DTT to SNAP determined the rate and duration of the recovery processes.


CONCLUSIONS
We suggest that NO can stimulate GABA release via GABA transporter reversal resulting from increased GABA levels in cytosol. The latter is reversible and appears to be due to S-nitrosylation of key proteins, which affect the energy status of the pre-synapse.


CONCLUSIONS
Our findings provide new insight into molecular mechanism(s) underlying the presynaptic action of nitric oxide on inhibitory neurotransmission.
Department of Neurochemistry, Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Leontovich Str. 9, Kyiv 01601, Ukraine. Electronic address: tas@biochem.kiev.ua.
Alla
A
Tarasenko
Department of Neurochemistry, Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Leontovich Str. 9, Kyiv 01601, Ukraine. Electronic address: olya_krupko@mail.ru.
Olga
O
Krupko
Department of Neurochemistry, Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Leontovich Str. 9, Kyiv 01601, Ukraine. Electronic address: ninahimm@biochem.kiev.ua.
Nina
N
Himmelreich
D000818:Animals; D002118:Calcium / Q000378:metabolism; D004229:Dithiothreitol / Q000494:pharmacology; D008297:Male; D053078:Membrane Potential, Mitochondrial; D009569:Nitric Oxide / Q000494:pharmacology; D020030:Nitric Oxide Donors / Q000494:pharmacology*; D017729:Presynaptic Terminals / Q000187:drug effects*; D051381:Rats; D017208:Rats, Wistar; D013572:Synaptic Vesicles / Q000378:metabolism; D005680:gamma-Aminobutyric Acid / Q000378:metabolism*
D020030:Nitric Oxide Donors; D009569:Nitric Oxide; D005680:gamma-Aminobutyric Acid; D002118:Calcium; D004229:Dithiothreitol
Exocytosis; Mitochondrial membrane potential; Nitric oxide; S-nitrosylation; Synaptic vesicle acidification; [(3)H]GABA release