Bradykinin-evoked Ca²⁺ mobilization in Madin Darby canine kidney cells

We studied the mechanisms underlying the bradykinin-evoked changes in intracellular calcium concentration ([Ca²⁺](i)) in Madin Darby canine kidney (MDCK) cells. Bradykinin evoked a [Ca²⁺](i) transient in a dose-dependent manner, measured by fura-2 fluorimetry and digital video imaging. The transient consisted of a rise and a decay and [Ca²⁺](i) returned to baseline without oscillations. External Ca²⁺ influx occurred, as demonstrated by Mn²⁺ quench and external Ca²⁺ removal measurements. Bradykinin acted by stimulating bradykinin B₂ receptors as evidenced by blockade by d-arginyl-l-arginlyl-l-prolyl-trans-4-hydroxy-l-prolylglycyl-3-(2-thienyl)-l-alanyl-l-seryl-d-1,2,3,4-tetrahydro-3-isoquinolinecarbonyl-l-(2α,3β,7aβ)-octahydro-1H-indole-2-carbonyl-l-arginine (HOE 140) but not by d-arginyl-l-arginlyl-l-prolyl-trans-4-hydroxy-l-prolylglycyl-3-(2-thienyl)-l-alanyl-l-seryl-d-1,2,3,4-tetrahydro-3-isoquinolinecarbonyl-l-(2α,3β,7aβ)-octahydro-1H-indole-2-carbonyl ([Des-Arg]HOE 140). The [Ca²⁺](i) signal was abolished by 1-(6-((17β-3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione (U73122) and partially inhibited by neomycin, implying mediation by phospholipase C. The transient was initiated by a release of Ca²⁺ from internal stores since it was abolished by pretreatment with thapsigargin or cyclopiazonic acid. The mobilization of the internal Ca²⁺ store subsequently triggered a 1-[β-[3-(4-methoxyphenyl)propoxy]-4-methoxyphenethyl]-1H-imidazole hydrochloride (SKF 96365)-insensitive Ca²⁺ entry. Pretreatment with carbonylcyanide m-chlorophynylhydrozone and gly-phe-β-naphthylamide did not alter the transient, thus excluding the participation of mitochondria and lysosomes. Efflux via Ca²⁺ pumps contributed to the decay of the transient. Efflux via Na⁺/Ca²⁺ exchange or sequestration by mitochondria and lysosomes was insignificant. The transient was blunted by the protein kinase C activator phorbol 12-myristate 13-acetate, and was enhanced by the protein kinase C inhibitors sphingosine and chelerythrine, the protein kinase A inhibitor 2,5-di-(t-butyl)-1,4-hydroquinone, N-[2-(p-bromocinnamylamino)ethyl]5-isoquinolinesulfonamide (H-89), the agent 8-(diethylamino)octyl 3,4,5-trimethoxybenzoate (TMB-8), and agents that elevated levels of 3',5'-cyclic guanosine monophosphate. The transient did not heterologously desensitize with that evoked by ATP, ADP or UTP. Copyright (C) 1998 Elsevier Science B.V.