Modulation of Bradykinin-Induced Inositol Trisphosphate Release in a Novel Neuroblastoma X Dorsal Root Ganglion Sensory Neuron Cell Line (F-11)

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Abstract
In the mouse neuroblastoma X dorsal root ganglion hybrid cell line F-l 1, bradykinin receptor stimulation induced the release of inositol-1,4,5-trisphosphate (IP3) and inositol-1,4-bisphosphate (IP2). Maximal stimulation of [2-3H]IP3 and [2-3H]IP2 release by bradykinin in the absence of LiCl occurred at 7 (or less) and 15 s, respectively, with average levels of 5.7- (IP3) and 3.4- (IP2) fold of control values. The EC50 for bradykinin was 33 ± 5 nM. IP3 and IP2 concentrations returned to basal levels ∼ 1 min after bradykinin addition. Bradykinin-induced IP3 release was blocked by several novel bradykinin analogues. In particular, [D-Arg0]-Hyp3-Thi58-[D-Phe7]-bradykinin [Hyp, hydroxypro-line; Thi, β-(2-thienyl)-L-alanine] blocked IP3 production in a dose-dependent fashion. Several of these analogues alone showed little or no agonist activity. The bradykinin receptor may be coupled to phospholipase C via a GTP-sensitive protein (Gi or Go), as preincubation for 18–20 h with pertussis toxin decreased IP3 concentrations by 45%. Bradykinin is also known to modulate the concentrations of other second messengers in neurons, increasing the concentrations of Ca2+, diacylglycerol (DG), and cyclic GMP and decreasing the concentration of cyclic AMP. These second messengers modulated bradykinin-dependent IP3 release to varying degrees. A23187, a Ca2+ ionophore, produced a 37% decrease in IP3 concentration. 12-O-Tetradecanoylphorbol-13-ace-tate, which mimics the effects of DG and activates protein kinase C, inhibited IP3 release by 80%. Dibutyryl cyclic GMP produced little or no inhibition of IP3. [D-Ala2,D-Leu5]Enkephalin (DADLE), an opioid peptide that decreases cyclic AMP concentrations, likewise had no effect. However, elevation of cyclic AMP concentrations by pros-taglandins I2 or E2 or forskolin inhibited IP3 formation in a dose-dependent fashion. This inhibition was reduced by DADLE in a naloxone-reversible manner, a result suggesting that the inhibition is a cyclic AMP-mediated effect. These results show that (a) bradykinin acutely stimulates IP3 release from F-11 cells in a transient fashion, (b) bradykinin-induced IP3 release may be subject to negative feedback control mediated through protein kinase C, (c) IP3 release is partially inhibited by pertussis toxin and by increases in cyclic AMP content, and (d) inhibition of bradykinin-induced IP3 release can be produced by several novel bradykinin analogues.