Possible involvement of calmodulin-dependent kinases in Ca(2+)-dependent enhancement of [3H]5-hydroxytryptamine uptake in rat cortex.
Effects of Ca2+ on [3H]5-hydroxytryptamine (5-HT) uptake into rat cortical synaptosomes were studied. The uptake was enhanced in the presence of Ca2+ in Krebs-Ringer medium and the uptake at 0.3-5 mM Ca2+ was 2.4-2.7 times greater than that observed in the absence of Ca2+. The maximal increase at the concentration of 1 mM Ca2+ was achieved after 2 min preincubation. Ca(2+)-dependent enhancement of the [3H]5-HT uptake reflected an increase in Vmax of the uptake process. However, Kd and Bmax values for [3H]paroxetine were not significantly changed in the presence of 1 mM Ca2+ compared with Ca(2+)-free condition. On the other hand, uptake was still enhanced after synaptosomes were washed with Ca(2+)-free after preincubation with 1 mM Ca2+. Staurosporine (a protein kinase C inhibitor) and wortmannin (a myosin light chain kinase inhibitor) did not affect Ca(2+)-dependent enhancement of the uptake, whereas 1-[N,O-bis(5-isoquinolinesulfonyl)-N-methyl-L-tyrosyl]-4-phenylpiperazin e (KN-62, an inhibitor of Ca2+ /calmodulin-dependent kinase II) and N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride (W-7, a calmodulin antagonist) significantly reduced it. Moreover, L-type, but not P- or N-type, voltage-dependent Ca(2+)-channel blockers suppressed enhancement of the uptake. These results indicate that Ca(2+)-dependent enhancement of [3H]5-HT uptake is mediated by activation of calmodulin-dependent protein kinases, suggesting a possibility of calmodulin-dependent regulation of in vivo 5-HT uptake.
Yura
A
A
Department of Pharmacology, Showa University School of Medicine, Tokyo, Japan.
Kiuchi
Y
Y
Uchikawa
T
T
Uchida
J
J
Yamazaki
K
K
Oguchi
K
K
D000818:Animals; D002118:Calcium / Q000494:pharmacology / Q000502:physiology*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000502:physiology*; D002147:Calmodulin / Q000037:antagonists & inhibitors; D002540:Cerebral Cortex / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008297:Male; D017374:Paroxetine / Q000378:metabolism; D047428:Protein Kinase Inhibitors; D051381:Rats; D017208:Rats, Wistar; D012701:Serotonin / Q000493:pharmacokinetics*; D013449:Sulfonamides / Q000494:pharmacology; D014316:Tritium
D002121:Calcium Channel Blockers; D002147:Calmodulin; D004791:Enzyme Inhibitors; D047428:Protein Kinase Inhibitors; D013449:Sulfonamides; D014316:Tritium; D012701:Serotonin; D017374:Paroxetine; C017967:W 7; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002118:Calcium