5-Hydroxytryptamine(1A) receptor/G(i)beta gamma stimulates mitogen-activated protein kinase via NAD(P)H oxidase and reactive oxygen species upstream of Src in Chinese hamster ovary fibroblasts

The hypothesis of this work is that the 'serotonin' or 5-hydroxytryptamine (5-HT)(1A) receptor, which activates the extracellular signal-regulated kin ase (ERK) through a G(1)beta gamma-mediated pathway, does so through the in termediate actions of reactive oxygen species (ROS). Five criteria were sho wn to support a key role for ROS in the activation of ERK by the 5-HT1A rec eptor. (1) Antioxidants inhibit activation of ERK by 5-HT. (2) Application of cysteine-reactive oxidant molecules activates ERK. (3) The 5-HT1A recept or alters cellular redox properties, and generates both superoxide and hydr ogen peroxide. (4) A specific ROS-producing enzyme [NAD(P)H oxidase] is inv olved in the activation of ERK. (5) There is specificity both in the effect s of various chemical oxidizers, and in the putative location of the ROS in the ERK activation pathway. We propose that NAD(P)H oxidase is located in the ERK activation pathway stimulated by the transfected 5-HT1A receptor in Chinese hamster ovary (CHO) cells downstream of G(1)beta gamma subunits an d upstream of or at the level of the non-receptor tyrosine kinase, Src. Mor eover, these experiments provide cofirmation that the transfected human 5-H T1A receptor induces the production of ROS (superoxide and hydrogen peroxid e) in CHO cells, and support the possibility that an NAD(P)H oxidase-like e nzyme might be involved in the 5-HT-mediated generation of both superoxide and hydrogen peroxide.