Perfused rat intestine for study of norepinephrine release

Previous preparations for studying the neuronal release of norepinephrine (NE) employed relatively large vessels, nonsanguinous perfusates, and the preloading of [3H]NE. To study the stimulated release of endogenous NE and the responses of true resistance vessels, we developed a rat intestine preparation that is pump perfused with canine red blood cells suspended in bicarbonate buffer with 6% albumin. In a pentobarbital-anesthetized rat, the duodenum, colon, and cecum are extirpated to isolate the ileum vascularly. After the superior mesenteric artery and vein are cannulated, the perivascular nerves are isolated to stimulate the postganglionic sympathetic fibers. To evaluate the preparation, we stimulated the sympathetic fibers at 1-10 Hz with supramaximal pulses. Resistance changes were assessed by monitoring perfusion pressure, and the concentration of NE was assayed in the venous effluent by the single-isotope radioenzymatic method. During nerve stimulation, the increases in both resistance and NE release rate were frequency dependent. Repetitions of electrical stimulation yielded reproducible frequency-response curves. Pretreatment with phentolamine (10 microM) abolished the resistance response and enhanced stimulated NE release, which roughly tripled at 10 Hz. Phentolamine at smaller doses (1 microM) eliminated the resistance responses to stimulation but did not enhance NE release. Cocaine alone (30 microM) increased base-line resistance and unstimulated NE release. After cocaine pretreatment, phentolamine at 1 microM enhanced the stimulated NE release rate. We conclude that the isolated rat intestine contains postsynaptic alpha-adrenoceptors that mediate vasconstriction and prejunctional alpha 2-adrenoceptors that mediate the inhibition of NE release. Thus the rat intestine is a responsive preparation for studying the release of endogenous NE and noradrenergic neurotransmission.