Extracellular pH, Ca(2+) influx, and response of vascular smooth muscle cells to 5-hydroxytryptamine.
OBJECTIVE
Cerebral vascular smooth muscle cells (VSMCs) contract on extracellular pH (pH(o)) increases and relax on pH(o) decreases. These changes in tone are believed to result from changes in [Ca(2+)](i), although the responsible mechanisms are not fully understood. VSMCs also contract in response to 5-hydroxytryptamine (5-HT), which increases [Ca(2+)](i) via both Ca(2+) release and influx. We hypothesized that examining effects of pH(o) decreases on 5-HT-induced [Ca(2+)](i) changes would allow us to identify mechanisms whereby pH(o) influences tone. Accordingly, we compared [Ca(2+)](i) increases in cerebral VSMCs, evoked by 5-HT, with increases evoked by increased pH(o) and examined 5-HT-dependent [Ca(2+)](i) increases at normal and decreased pH(o).


METHODS
We monitored [Ca(2+)](i,), using the Ca(2+)-sensitive dye fura 2, in cultured rat cerebral VSMCs obtained by enzymatic digestion of middle cerebral arteries and their branches (passages 1 to 3) grown on glass coverslips and superfused with physiological saline.


RESULTS
Increasing pH(o) from 7.3 to 7.8 increased [Ca(2+)](i), and these increases were prevented in Ca(2+)-free solutions. Decreasing pH(o) from 7.3 to 6.9 did not alter [Ca(2+)](i) unless [Ca(2+)](i) was first raised by treatment with 5-HT (10 micromol/L). 5-HT resulted in biphasic [Ca(2+)](i) increases characterized by transient peaks blocked by the Ca(2+)-ATPase inhibitor thapsigargin (10 nmol/L) and prolonged plateaus blocked by the Ca(2+) channel blocker Ni(2+) (1 mmol/L). Acidification did not alter the transient peaks but significantly reduced 5-HT-induced Ca(2+) influx.


CONCLUSIONS
We conclude that increasing pH(o) induces Ca(2+) influx in rat cerebral VSMCs and decreasing pH(o) inhibits 5-HT-stimulated Ca(2+) entry but not intracellular Ca(2+) release.
Nazarov
V
V
Departments of Pediatrics, Yale University School of Medicine, New Haven, Connecticut, USA.
Aquino-DeJesus
J
J
Apkon
M
M
D000143:Acids / Q000494:pharmacology; D000468:Alkalies / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D002560:Cerebrovascular Circulation / Q000187:drug effects; D005110:Extracellular Space / Q000378:metabolism*; D005456:Fluorescent Dyes; D016257:Fura-2; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012701:Serotonin / Q000378:metabolism* / Q000494:pharmacology
D000143:Acids; D000468:Alkalies; D005456:Fluorescent Dyes; D012701:Serotonin; D002118:Calcium; D016257:Fura-2