Involvement of protein kinases on the upregulation of endothelin receptors in rat basilar and mesenteric arteries

Endothelin(B) (ETB) receptors are upregulated in experimental stroke or after 24 hrs of organ culture. This upregulation is manifested both as stronger contraction and as an increase in ETB receptor messenger RNA (mRNA) levels. The present study was designed to evaluate the importance of protein kinases (c-Jun N-terminal kinase [JNK], protein kinase C [PKC], and extracellular signal-regulated kinase [ERK1/2]) in ETB receptor upregulation after organ culture. Rat basilar and mesenteric arteries were incubated for 24 hrs in Dulbecco's modified Eagle's medium (DMEM) with or without the PKC inhibitor, RO31-7549; the ERK1/2 inhibitor, SB386023; or the JNK inhibitor, SP600125, added 3, 6, or 12 hrs after initiation of incubation. Subsequently, vessel segments were mounted in myographs and the contractile responses to ET-1 and sarafotoxin 6c were studied. The ETB and ETA receptor mRNA levels were determined with a real-time polymerase chain reaction (PCR). The cellular localization and protein level of ETB receptors were evaluated by immunohistochemistry. The PKC and ERK1/2 inhibitors attenuated the contraction induced by S6c in the basilar arteries more than in the mesenteric arteries. The efficiency of the inhibitors was proportional to the incubation time. Real-time PCR showed a decrease in the ETB receptor mRNA levels in arteries treated with PKC or ERK inhibitors. The JNK inhibitor had a significant inhibitory effect on ETB receptor upregulation in the basilar arteries. Immunohistochemistry revealed that the ETB receptor upregulation occured in the smooth-muscle cells and that it had the same pattern as in the quantitative PCR. Our results show that the PKC, ERK1/2, and JNK are more important for the upregulation of contractile ETB receptors in cerebral arteries compared with mesenteric arteries. ERK1/2 seems to be more important for the ETB receptor upregulation, as compared with PKC and JNK. The evaluation of the time dependency suggests that the phenomenon can be reversed even after its initiation.