Facilitation of dopamine and acetylcholine release by intermittent hypoxia in PC12 cells: involvement of calcium and reactive oxygen species.
We have investigated the effects of preconditioning pheochromocytoma (PC12) cells with intermittent hypoxia (IH) on transmitter release during acute hypoxia. Cell cultures were exposed to either alternating cycles of hypoxia (1% O(2) + 5% CO(2); 30 s/cycle) and normoxia (21% O(2) + 5% CO(2); 3 min/cycle) for 15 or 60 cycles or normoxia alone (control) for similar durations. Control and IH cells were challenged with either hyperoxia (basal release) or acute hypoxia (Po(2) of approximately 35 Torr) for 5 min, and the amounts of dopamine (DA) and acetylcholine (ACh) released in the medium were determined by HPLC combined with electrochemical detection. Hypoxia augmented DA (approximately 80%) but not ACh release in naive cells, whereas, in IH-conditioned cells, it further enhanced DA release (ranging from 120 to approximately 145%) and facilitated ACh release (approximately 30%). Hypoxia-evoked augmentation of transmitter release was not seen in cells conditioned with sustained hypoxia. IH-induced increase in DA but not IH-induced ACh release during hypoxia was partially inhibited by cadmium chloride (100 microM), a voltage-gated Ca(2+) channel blocker. By contrast, 2-aminoethoxydiphenylborate (75 microM), a blocker of inositol 1,4,5-trisphosphate (IP(3)) receptors, and N-acetyl-L-cysteine (300 microM), a potent scavenger of reactive oxygen species, either attenuated or abolished IH-evoked augmentation of transmitter release during hypoxia. Together, the above results demonstrate that IH conditioning increases hypoxia-evoked neurotransmitter release from PC12 cells via mechanisms involving mobilization of Ca(2+) from intracellular stores through activation of IP(3) receptors. Our findings also suggest that oxidative stress plays a central role in IH-induced augmentation of transmitter release from PC12 cells during acute hypoxia.
Kim
Dong-Kyu
DK
Department of Biochemistry, Case Western Reserve University, Cleveland, OH 44106-4935, USA.
Natarajan
Niranjana
N
Prabhakar
Nanduri R
NR
Kumar
Ganesh K
GK
D000109:Acetylcholine / Q000378:metabolism*; D000818:Animals; D019256:Cadmium Chloride / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D015687:Cell Hypoxia / Q000187:drug effects / Q000502:physiology; D004298:Dopamine / Q000378:metabolism*; D016716:PC12 Cells; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism*
D017382:Reactive Oxygen Species; D019256:Cadmium Chloride; D000109:Acetylcholine; D002118:Calcium; D004298:Dopamine