Alpha 1-adrenoceptor activation increases ecto-5'-nucleotidase activity and adenosine release in rat cardiomyocytes by activating protein kinase C.
BACKGROUND
Adenosine is an important regulator of many cardiac functions and is synthesized primarily by ecto- and cytosolic 5'-nucleotidase. We have previously reported that alpha 1-adrenoceptor blockade attenuates adenosine release from ischemic myocardium, raising the possibility that alpha 1-adrenoceptor activation activates 5'-nucleotidase. This study tested whether activation of protein kinase C by alpha 1-adrenoceptor activation increases 5'-nucleotidase activity and augments adenosine release.


RESULTS
Cardiomyocytes were isolated from adult male Wistar rats and suspended in modified HEPES-Tyrode's buffer solution. After stabilization, the cardiomyocytes were incubated with and without an exposure to norepinephrine (10(-9) to 10(-5) mol/L) while being treated with propranolol and yohimbine or with and without an exposure to methoxamine (10(-9) to 10(-5) mol/L). Ecto-5'-nucleotidase activity was increased by norepinephrine and methoxamine during 30 minutes in a dose-dependent manner, whereas cytosolic 5'-nucleotidase was not activated. These increases in ecto-5'-nucleotidase activity were inhibited by GF109203X, an inhibitor of protein kinase C, and mimicked by phorbol 12-myristate 13-acetate (PMA), an activator of protein kinase C. The increase in ecto-5'-nucleotidase was not prevented by cycloheximide. When ecto-5'-nucleotidase activity increased, adenosine release was augmented in methoxamine- and PMA-treated cardiomyocytes (1299 +/- 252% and 1372 +/- 149%, respectively) compared with the untreated group (578 +/- 26%). The increase in adenosine release was blunted by GF109203X and alpha, beta-methyleneadenosine 5'-diphosphate, an inhibitor of ecto-5'-nucleotidase.


CONCLUSIONS
Thus, we conclude that alpha 1-adrenoceptor-mediated increases in ecto-5'-nucleotidase activity are attributed to activation of protein kinase C in rat cardiomyocytes.
Kitakaze
M
M
First Department of Medicine, Osaka University School of Medicine, Suita, Japan.
Hori
M
M
Morioka
T
T
Minamino
T
T
Takashima
S
S
Okazaki
Y
Y
Node
K
K
Komamura
K
K
Iwakura
K
K
Itoh
T
T
D015720:5'-Nucleotidase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000241:Adenosine / Q000378:metabolism*; D000244:Adenosine Diphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D000249:Adenosine Monophosphate / Q000378:metabolism; D000818:Animals; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D066298:In Vitro Techniques; D007211:Indoles / Q000494:pharmacology; D008297:Male; D008301:Maleimides / Q000494:pharmacology; D008729:Methoxamine / Q000494:pharmacology; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D009638:Norepinephrine / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D018340:Receptors, Adrenergic, alpha-1 / Q000187:drug effects / Q000502:physiology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D007211:Indoles; D008301:Maleimides; D018340:Receptors, Adrenergic, alpha-1; C523965:alpha,beta-methyleneadenosine 5'-diphosphate; D000249:Adenosine Monophosphate; D000244:Adenosine Diphosphate; D011493:Protein Kinase C; D015720:5'-Nucleotidase; D008729:Methoxamine; D000241:Adenosine; C070515:bisindolylmaleimide I; D013755:Tetradecanoylphorbol Acetate; D009638:Norepinephrine