Activation of recombinant human TRPV1 receptors expressed in SH-SY5Y human neuroblastoma cells increases [Ca2+]i, initiates neurotransmitter release and promotes delayed cell death

Abstract

The transient receptor potential (TRP) vanilloid receptor subtype 1 (TRPV1) is a ligand-gated, Ca2+-permeable ion channel in the TRP superfamily of channels. We report the establishment of the first neuronal model expressing recombinant human TRPV1 (SH-SY5YhTRPV1). SH-SY5Y human neuroblastoma cells were stably transfected with hTRPV1 using the Amaxa Biosystem (hTRPV1 in pIREShyg2 with hygromycin selection). Capsaicin, olvanil, resiniferatoxin and the endocannabinoid anandamide increased [Ca2+]i with potency (EC50) values of 2.9 nmol/L, 34.7 nmol/L, 0.9 nmol/L and 4.6 μmol/L, respectively. The putative endovanilloid N-arachidonoyl-dopamine increased [Ca2+]i but this response did not reach a maximum. Capsaicin, anandamide, resiniferatoxin and olvanil mediated increases in [Ca2+]i were inhibited by the TRPV1 antagonists capsazepine and iodo-resiniferatoxin with potencies (KB) of ∼70 nmol/L and 2 nmol/L, respectively. Capsaicin stimulated the release of pre-labelled [3H]noradrenaline from monolayers of SH-SY5Y hTRPV1 cells with an EC50 of 0.6 nmol/L indicating amplification between [Ca2+]i and release. In a perfusion system, we simultaneously measured [3H]noradrenaline release and [Ca2+]i and observed that increased [Ca2+] i preceded transmitter release. Capsaicin treatment also produced a cytotoxic response (EC50 155 nmol/L) that was antagonist-sensitive and mirrored the [Ca2+]I response. This model displays pharmacology consistent with TRPV1 heterologously expressed in standard non-neuronal cells and native neuronal cultures. The advantage of SH-SY5Y hTRPV1 is the ability of hTRPV1 to couple to neuronal biochemical machinery and produce large quantities of cells. © 2007 The Authors.