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

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.