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

Abstract

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 [Ca 2+ ] i was examined in dissociated rat spinal cord neurons by fura 2 microfluorimetry. The neurons investigated were choline acetyltransferasepositive and had morphological features of motoneurons. U-II induced [Ca 2+ ] i increases in these neurons with a threshold of 10 )9 M, and a maximal effect at 10 )6 M with an estimated EC 50 of 6.2 · 10 )9 M. The [Ca 2+ ] i increase induced by U-II was mainly caused by Ca 2+ influx from extracellular space, as the response was markedly attenuated in a Ca 2+ -free medium. Omega-conotoxin GVIA (10 )7 M), a N-type Ca 2+ channel blocker, largely inhibited these increases, whereas the P/Q Ca 2+ channel blocker, omega-conotoxin GVIIC (10 )7 M) and the L-type Ca 2+ channel blocker, verapamil (10 )5 M) had minimal effects. Down-regulation of protein kinase C by 4-aphorbol 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 (3 · 10 )5 M) did not modify U-II-induced [Ca 2+ ] i increases. In contrast, protein kinase A inhibitors KT5720 (10 )6 M) and Rp-cAMPS (3 · 10 )5 M) reduced the response to 25 ± 3% and 42 ± 8%, respectively. Present results demonstrate that U-II modulates [Ca 2+ ] i in rat spinal cord neurons via protein kinase A cascade.

MOLECULAR EVOLUTION OF GPCRS: Somatostatin/urotensin II receptors

“…The presence of UII receptor on cholinergic nerve terminals, in part originating from the recurrent motoneuron axon collaterals, in mouse spinal cord has given rise to the idea that UII may regulate motoneuron activity in an autocrine/paracrine manner (Bruzzone et al 2010). Supporting this view, functional UII receptors have been found to be coupled to calcium mobilization in dissociated rat motor neurons (Filipeanu et al 2002). A presynaptic role at the neuromuscular junction is also likely, as UII immunoreactivity is detected in muscle motor end plate regions coexisting with synaptophysin immunolabeling and because UII promotes acetylcholine release (Brailoiu et al 2003, Bruzzone et al 2010.…”

Nicotinic Acid Adenine Dinucleotide Phosphate Potentiates Neurite Outgrowth

“…Measurement of Cytosolic Ca 2ϩ Concentration-Cytosolic Ca 2ϩ concentration measurements were performed as described previously (27). Briefly, freshly dissociated neurons (2 h after isolation) were loaded with the fluorescent Ca 2ϩ indicator Fura-2 by incubation of the cells in Hank's balanced salt solution supplemented with 3 M Fura-2/AM for 45 min and Hank's balanced salt solution alone for an additional 15-60 min (to allow de-esterification of the dye).…”

International Union of Basic and Clinical Pharmacology. XCII. Urotensin II, Urotensin II–Related Peptide, and Their Receptor: From Structure to Function

“…In endothelium-denuded rat aorta, protein kinase C mediates the synergistic action of UII and angiotensin II . In addition, in rat spinal cord cholinergic neurons, UII causes calcium influx from the extracellular space via N-type Ca 2+ channels, and this effect is mediated through the protein kinase A pathway (Filipeanu et al, 2002), whereas in arterial smooth muscle cells, UII stimulates Ca 2+ influx via L-type Ca 2+ channels (Sauzeau et al, 2001) indicating that, depending on the cell type, UII-induced Ca 2+ entry occurs through various types of voltage-operated Ca 2+ channels. Of note, UII provokes membrane depolarization in cholinergic neurons from the ventral tegmentum that is likely involved in the UII-evoked control of rapid eye movements (de Lecea and Bourgin, 2008).…”