Neuropeptide Y induced increase of cytosolic and nuclear Ca2+in heart and vascular smooth muscle cells

Abstract

It was reported that neuropeptide Y (NPY) affects cardiac and vascular smooth muscle (VSM) function probably by increasing intracellular Ca2+. In this study, using fura-2 microfluorometry and fluo-3 confocal microscopy techniques for intracellular Ca2+measurement, we attempted to verify whether the action of NPY receptor's stimulation in heart and VSM cells modulates intracellular Ca2+and whether this effect is mediated via the Y1receptor type. Using spontaneously contracting single ventricular heart cells of 10-day-old embryonic chicks and the fluo-3 confocal microscopy Ca2+measurement technique to localize cytosolic ([Ca]c) and nuclear ([Ca]n) free Ca2+level and distribution, 10-10M of human (h) NPY significantly (P < 0.05) increased the frequency of cytosolic and nuclear Ca2+transients during spontaneous contraction. Increasing the concentration of hNPY (10-9M) did not further increase the frequency of Ca2+transients. The L-type Ca2+channel blocker, nifedipine (10-5M), significantly (P < 0.001) blocked the spontaneous rise of intracellular Ca2+in the absence and presence of hNPY (10-10and 10-9M). However, the selective Y1receptor antagonist, BIBP3226 (10-6M), significantly decreased the hNPY-induced (10-10and 10-9M) increase in the frequency of Ca2+transients back to near the control level (P < 0.05). In resting nonworking heart and human aortic VSM cells, hNPY induced a dose-dependent sustained increase of basal resting intracellular Ca2+with an EC50near 10-9M. This sustained increase was cytosolic and nuclear and was completely blocked by the Ca2+chelator EGTA, and was significantly decreased by the Y1receptor antagonist BIBP3226 in both heart (P < 0.05) and VSM (P < 0.01) cells. These results strongly suggest that NPY stimulates the resting basal steady-state Ca2+influx through the sarcolemma and induces sustained increases of cytosolic and nuclear calcium, in good part, via the activation of the sarcolemma membrane Y1receptor type in both resting heart and VSM cells. In addition, NPY also increased the frequency of Ca2+transients during spontaneous contraction of heart cells mainly via the activation of the Y1receptor type, which may explain in part the active cardiovascular action of this peptide.Key words: heart, vascular smooth muscle, neuropeptide Y, BIBP3226, calcium, nucleus.