Purinergic Component Involved in Long Vasodilatory Reflex in the Guinea Pig Small Intestine
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Submucosal arterioles in the small intestine are the main point of control for gastrointestinal (GI) circulation as they are the final resistance vessels feeding the highly perfused mucosal layer. Ischemia can lead to pathophysiology of a variety of GI tissues. In chronic intestinal inflammation, alterations in blood flow have been purported to be involved in disease etiology. The aim of this study was to characterize purinergic neurotransmitter pathways involved in physiological submucosal arteriole diameter control by the enteric nervous system long vasodilatory reflex (LVD) and to establish a protocol to determine how inflammatory neural changes affect vasodilation in the small intestine. Following euthanasia, segments of small intestine were harvested from adult male guinea pigs and changes in nerve stimulated small intestine submucosal arteriole diameter were identified using videomicroscopy techniques; vessels were preconstricted and nicotinic cholinergic transmission was blocked with hexamethonium. Purinergic receptor antagonists were applied. Immunohistochemical analysis was conducted to identify P2Y1 receptors localization. In a subset of experiments sensory neuronal excitability was initiated using phorbol dibutyrate (PDBu) shown previously to induce hyperexcitability in the sensory neurons similar to changes found in intestinal inflammation. In these experiments, intestinal segments were placed into a novel dual chamber bath separated into two portions and PDBu was applied unilaterally. Blood vessel vasodilation was either abolished or decreased by both suramin (100 μM, n=6), a non-specific P2 purinergic antagonist. MRS 2179 (10 μM, n=5), a P2Y1 specific antagonist, also decreased vasodilation, which suggests that there is a purinergic neurotransmission component to the LVD mediated by P2 receptors, including the P2Y1 subtype. Immunohistochemistry identified P2Y1 receptor staining that was uniformly punctated in both the myenteric and submucosal plexuses but specific neuronal locations of the receptor could not be identified. Nerve stimulated vasodilation was not altered by application of PDBu suggesting that neuronal hypersensitivity did not modify vessel dilation. Taken together these data suggest that purinergic receptor pathways contribute to the LVD reflex under normal conditions however more experiments are still required to fully elucidate how these pathways are affected /altered by intestinal inflammation.