Newoscience Vol. 62, No. I, pp. 125-143, 1994 Elsevier Science Ltd Copyright 0 1994 IBRO Printed in Great Britain. All rights reserved 0306-4522!94 $7.00 + 0.00 0306-4522(94)EO156-X zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQ EVIDENCE FOR WIDESPREAD AFFERENTS TO BARRINGTO~S NUCLEUS, A BRAINSTEM REGION RICH IN CORTICOTROPIN-RELEASING HORMONE NEURONS R. J. VALEN~NO,* kl. E. PAGE, P.-H. LUPPI, Y. ZHU, E. VAN BOCKSTAELE and G. ASTON-JONES Division of Behavioral Neurobiology, Department of Mental Health Sciences, Hahnemann University, Broad and Vine Sts, Philadelphia, PA 19102-1192, U.S.A. Abstract-Supraspinal a&rents to the pentine micturition center, Barrington’s nucleus, were investigated in the rat by visualization of the retrograde tracer, cholera-toxin subunit B, in neurons following iontophoretic injection into Barrington’s nucleus. Tissue sections from five rats with injections primarily localized in Barrington’s nucleus revealed numerous retrogradely labeled neurons throughout all rostrocaudal levels of the ~~aqu~uctal gray (particularly its ventrolatemi division), in the lateral hypothalamic area (particularly medial to the fornix), and in the medial preoptic nucleus. Retrogfadely labeled neurons were also consistently found in the nucleusof the solitary tract, in the vicinity of the lateral reticular nucleus, nucleus paragigantocellularis, parabrachial nucleus, KSlliker-Fuse nucleus, cuneiform nucleus, raphe nucleus and zona incerta. In the hypothalamus, in addition to the perifornical region, retrogradely labeled neurons were found in all cases in the tuberomammill~ nucleus, premammilla~ nucleus, dorsal hy~thalamic area, ventrom~ial h~othalamic nucleus, and the paravent~c~ar nucleus. At more rostra1 levels, in addition to the medial preoplic area, retrogradely labeled neurons were seen in the bed nucleus of the stria terminalis and in a region just lateral to the supraoptic nucleus near the medial amygdaloid nucleus. Retrogradely labeled neurons were also observed in the motor, insular, and infralimbic cortices. Injections of anterograde tracers (cholera-toxin subunit B or zyxwvutsrqponmlkjihgfedcbaZ Phuseolus vulgaris leucoagglutinin) into the K~iliker-Fur nucleus, the ventrola~ral ~~aqu~u~tal gray, lateral hypothala- mic area, or medial preoptic area, resulted in fiber labeling within Barrington’s nucleus, confirming the retrograde tracing studies. As previously reported, numerous neurons in Barrington’s nucleus were immunoreactive for corticotropin-releasing hormone. Double-labelingstudies revealed afferent fibers from the periaqueductal gray and lateral hypothalamic area overlapping the corticotropin-releasing hormone- immunorea~ive neurons of Bar~n~on’s nucleus, and in some cases anterogmdely labeled fibers with varcosities appeared to target these neurons. The present results suggest that Barrington’s nucleus in the rat receives neuronal inputs from brainstem nuclei as well as from forebrain limbic structures including hypothalamic nuclei, the medial preoptic nucleus, and cortical areas involved in fluid balance or blood pressure regulation. In light of the role of Barrington’s nucleus in mi~urition, the integration of these various inputs may be important for co-ordinating urinary fun&on with fluid and cardiovascular homeostasis. Additionally, as neurons in Barrington’s nucleus are immunoreactive for the stress-related neurohormone, corticotropin-releasing hormone, these diverse inputs may regulate stress-related functions of this nucleus. The discovery of a region in the dorsolateral pons that regulates micturition-‘” led to numerous electro- physiological,‘4”u~35 anatomical,‘*~‘~~is~3~43 and le- sion33.3s studies to better define this region. These studies indicated that a small group of neurons lying just ventromedial to the rostra1 pole of the nucleus locus coeruleus (LC), termed Bar~ngton’s nucleus, ‘To whom correspondence should be addressed. Abbreviations: CRH, corticotropin-releasing hormone: CTb, cholera-toxin subunit B; BBST, bicarbonate- buffered saline with Triton X-100; DAB, 3,3’-di- aminobenzidine 4 HCI; DBH, dopamine /?-hydroxylase; HRP, horseradish peroxidase; LC, nucleus locus coeruleus: Me% mesenceohalic trieeminal nucleus: PAG, periaqueductal’ gray; P&, phosphate-buffered ‘saline: PHA-L, Pb~eoi~ vuigaris leucoagglu~in; PBST, phos- p~t5buffe~ saline with Triton X-100; Sol, nucleus of the solitary tract. was integral to the control of micturition. Electrical or chemical stimulation of this region in rats or cats elicits bladder cont~ction and increases discharge of bladder postganglionic nerves.14,20.‘3.35 Conversely, lesions of this nucleus33~3s or application of opiates inhibit micturition.44Consistent with this, anatomical studies utili~ng anterograde tracers and degeneration have demonstrated that the neurons of Barrington’s nucleus project to the intermediolateral column of the sacral spinal cord in the region of preganglionic neurons that innervate the bladder.‘2.‘s,‘a Similariy, injection of retrograde tracers into the intermediolat- era1 column of the sacral spinal cord labels neurons in the area corresponding to Barrington’s nu- cleus.‘2*‘8’32’43 Taken together, these electrophysiologi- cal and anatomical studies implicate Barrington’s nucleus in micturition. 125