Urotensin-II regulates intracellular calcium in dissociated rat spinal cord neurons

Urotensin-II (U-II), a peptide with multiple vascular effects, is detected in cholinergic neurons of the rat brainstem and spinal cord. Here, the effects of U-II on [Ca2+](i) was examined in dissociated rat spinal cord neurons by fura 2 microfluorimetry. The neurons investigated were choline acetyltransferase-positive and had morphological features of motoneurons. U-II induced [Ca2+](i) increases in these neurons with a threshold of 10(-9) M, and a maximal effect at 10(-6) M with an estimated EC50 of 6.2x10(-9) M. The [Ca2+](i) increase induced by U-II was mainly caused by Ca2+ influx from extracellular space, as the response was markedly attenuated in a Ca2+-free medium. Omega-conotoxin GVIA (10(-7) M), a N-type Ca2+ channel blocker, largely inhibited these increases, whereas the P/Q Ca2+ channel blocker, omega-conotoxin GVIIC (10(-7) M) and the L-type Ca2+ channel blocker, verapamil (10(-5) M) had minimal effects. Down-regulation of protein kinase C by 4-alpha-phorbol 12-myristate 13-acetate (10(-6) M) or enzyme inhibition using the specific inhibitor bisindolylmaleimide I (10(-6) M) did not inhibit the observed effects. Similarly, inhibition of protein kinase G with KT5823 (10(-6) M) or Rp-8-pCPT-cGMPS (3x10(-5) M) did not modify U-II-induced [Ca2+](i) increases. In contrast, protein kinase A inhibitors KT5720 (10(-6) M) and Rp-cAMPS (3x10(-5) M) reduced the response to 25+/-3% and 42+/-8%, respectively. Present results demonstrate that U-II modulates [Ca2+](i) in rat spinal cord neurons via protein kinase A cascade.