Functional expression of bradykinin B1 and B2 receptors in neonatal rat trigeminal ganglion neurons

Abstract

Bradykinin (BK) and its receptors (B1 and B2 receptors) play important roles in inflammatory nociception. However, the patterns of expression and physiological/pathological functions of B1 and B2 receptors in trigeminal ganglion (TG) neurons remain to be fully elucidated. We investigated the functional expression of BK receptors in rat TG neurons. We observed intense immunoreactivity of B2 receptors in TG neurons, while B1 receptors showed weak immunoreactivity. Expression of the B2 receptor colocalized with immunoreactivities against the pan-neuronal marker, neurofilament H, substance P, isolectin B4, and tropomyosin receptor kinase A antibodies. Both in the presence and absence of extracellular Ca2+ ([Ca2+]o), BK application increased the concentration of intracellular free Ca2+ ([Ca2+]i). The amplitudes of BK-induced [Ca2+]i increase in the absence of [Ca2+]o were significantly smaller than those in the presence of Ca2+. In the absence of [Ca2+]o, BK-induced [Ca2+]i increases were sensitive to B2 receptor antagonists, but not to a B1 receptor antagonist. However, B1 receptor agonist, Lys-[Des-Arg9]BK, transiently increased [Ca2+]i in primary cultured TG neurons, and these increases were sensitive to a B1 receptor antagonist in the presence of [Ca2+]o. These results indicated that B2 receptors were constitutively expressed and their activation induced the mobilization of [Ca2+]i from intracellular stores with partial Ca2+ influx by BK. Although constitutive B1 receptor expression could not be clearly observed immunohistochemically in the TG cryosection, cultured TG neurons functionally expressed B1 receptors, suggesting that both B1 and B2 receptors involve pathological and physiological nociceptive functions.

Intracellular Ca2+ mobilization pathway via bradykinin B1 receptor activation in rat trigeminal ganglion neurons

“…We therefore concluded that B 1 receptors in TG neurons, similar to those elsewhere in the brain, show high selectivity for Lys-[Des-Arg 9 ]BK [3]. In addition, the activation of B 2 receptors induced both the influx of Ca 2+ from the extracellular medium and the release of Ca 2+ from intracellular Ca 2+ stores [3]. However, the intracellular signaling pathway by which Lys-[Des-Arg 9 ]BK induces Ca 2+ mobilization in response to the activation of B 1 receptors had not yet been fully elucidated.…”

“…In a previous study [3], we demonstrated functional expression of B 1 and B 2 receptors in TG neurons, observing that BK elicited increases in intracellular free Ca 2+ concentration ([Ca 2+ ] i ) that were inhibited by B 2 receptor antagonists, but not by B 1 receptor antagonists, whereas application of Lys-[Des-Arg 9 ] BK induced increases in [Ca 2+ ] i that were sensitive to a B 1 receptor antagonist. We therefore concluded that B 1 receptors in TG neurons, similar to those elsewhere in the brain, show high selectivity for Lys-[Des-Arg 9 ]BK [3]. In addition, the activation of B 2 receptors induced both the influx of Ca 2+ from the extracellular medium and the release of Ca 2+ from intracellular Ca 2+ stores [3].…”

“…A solution containing a high concentration of extracellular K + (91 mM NaCl, 50 mM KCl, 2.0 mM CaCl 2 , 0.5 mM MgCl 2 , 0.44 mM KH 2 PO 4 , 0.34 mM Na 2 HPO 4 , 4.17 mM NaHCO 3 , 5.55 mM glucose; pH 7.4) was used to distinguish TG neurons from glial cells through activation of depolarization-induced increases in [Ca 2+ ] i in the neurons. The endogenous potent and highly selective bradykinin B 1 receptor agonist Lys-[Des-Arg 9 ]BK [3], the sarco/endoplasmic reticulum Ca 2+ -ATPase (SERCA) inhibitor cyclopiazonic acid (CPA, 100 nM [5]), the ryanodine receptor inhibitor dantrolene (sodium salt, 1 μM [5,6]), the phosphodiesterase (PDE) inhibitor isobutylmethylxanthine (IBMX, 50 μM [5]), the phospholipase C inhibitor U73122 (100 nM [7]) and the adenylyl cyclase inhibitor SQ22536 (1 μM [5,8]) were obtained from Tocris Bioscience (Bristol, UK). Xestospongin C [9], which antagonizes the calciumreleasing action of IP 3 at the receptor level without interacting with the IP 3 -binding site, was purchased from Wako Pure Chemical Industries, Ltd. (Osaka, Japan).…”

“…B 1 receptors have been suggested as an attractive target for the control of neuropathic pain [2]. In a previous study [3], we demonstrated functional expression of B 1 and B 2 receptors in TG neurons, observing that BK elicited increases in intracellular free Ca 2+ concentration ([Ca 2+ ] i ) that were inhibited by B 2 receptor antagonists, but not by B 1 receptor antagonists, whereas application of Lys-[Des-Arg 9 ] BK induced increases in [Ca 2+ ] i that were sensitive to a B 1 receptor antagonist. We therefore concluded that B 1 receptors in TG neurons, similar to those elsewhere in the brain, show high selectivity for Lys-[Des-Arg 9 ]BK [3].…”

“…Neuropathic pain, which is mediated by both B 1 and B 2 receptor activation in the orofacial area, is often induced by injuries to trigeminal ganglion (TG) neurons or glial cells [2][3][4]. B 1 receptors have been suggested as an attractive target for the control of neuropathic pain [2].…”