Sphingomyelinase and ceramide analogs induce contraction and rises in [Ca2+](i) in canine cerebral vascular muscle

Studies were designed to investigate effects of neutral sphingomyelinase (N -SMase) and ceramide analogs as well as phosphorylcholine on vascular tone and Ca2+ mobilization in isolated canine cerebral arterial smooth muscle. N -SMase (0.001-0.1 U/ml) provoked a gradual but sustained vasoconstriction o f arterial rings in a concentration-related manner that was endothelium ind ependent. Incubation of denuded arterial rings in Ca2+-free medium or pretr eatment with verapamil in extracellular Ca2+ resulted in a reduction of the N-SMase-evoked constriction. Exposure of arterial rings to 1,2-bis(2-amino phenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA)-AM did not, however, resu lt in a reduction of N-SMase-induced constriction. Both staurosporine and b isindolymaleimide I attenuated N-SMase-induced contractions to 66% and 72% of control, respectively. N-SMase caused gradual and sustained rises in int racellular Ca2+ concentration ([Ca2+](i) in primary cultured cerebral vascu lar smooth muscle cells. Pretreatment of these cultured cells with nimodipi ne and verapamil caused a steady decline in N-SMase-induced rises in [Ca2+] (i). Exposure of the cells to Ca2+-free solution reversed the [Ca2+](i)-ind uced rise triggered by N-SMase to the resting baseline. Both C-8 and C-16 c eramide (10(-9)-10(-6) M), but not phosphorylcholine, constricted denuded c anine arterial rings in a concentration-related manner and elevated [Ca2+]( i). Our results suggest that the sphingomyelin-signaling pathway, via a pro bable release of ceramide molecules, may play an important role in regulati on of cerebral arterial wall tone.