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NEUROREGULATION AND MOTILITY

Upregulation of cyclooxygenase-2 and thromboxane A₂ production mediate the action of tumor necrosis factor- in isolated rat myenteric ganglia

Institute for Veterinary Physiology, University Giessen, Giessen, Germany

Submitted 18 January 2005 ; accepted in final form 2 May 2005

Intact myenteric ganglia from 4- to 10-day-old rats were isolated from the small intestine. The preparations were cultured overnight, and drugs were applied within this time frame (20 h). Whole cell patch-clamp technique was used to measure basal membrane potential and carbachol-induced depolarization at neurons within these ganglia. Pretreatment with TNF- (100 ng/ml) hyperpolarized the membrane (from –31.0 ± 2.7 mV under control conditions to –61.2 ± 3.2 mV in the presence of the cytokine) and potentiated the depolarization induced by carbachol (from 5.2 ± 0.7 mV under control conditions to 27.5 ± 2.0 mV in the presence of the cytokine). These effects were mimicked by carbocyclic thromboxane A₂ (10^–6 mol/l), a stable thromboxane A₂ agonist. The TNF- action was inhibited by 1-benzylimidazole (2 x 10^–4 mol/l), a thromboxane synthase inhibitor, and BAY U 3405 (5 x 10^–4 mol/l), an inhibitor of thromboxane receptors. Measurements of thromboxane production in the supernatant of the culture revealed an increased concentration of thromboxane B₂, the stable metabolite of thromboxane A₂, after exposure to TNF-. Immuncytochemical staining for cyclooxygenase-2 (COX-2) and the neuronal marker microtubule-associating protein-2 revealed an upregulation of COX-2 in myenteric neurons after exposure to the cytokine. These results demonstrate the involvement of COX-2 and the subsequent production of thromboxane A₂ in the presence of TNF-.

COX-2; membrane potential

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