Brain Research, 587 (1992) 269-275 © 1992 Elsevier Science Publishers B.V. All rights reserved 0006-8993/92/$05.00 269 BRES 17964 Evidence that/3-endorphin is synthesized in cells in the m cleus tractus solitarius: detection of POMC mRNA David M. Bronstein, Martin K.-H. Schafer *, Stanley J. Watson and Huda Akil Mental Health Research h~stitute, Unirersity of Michigan, Ann Arbor, Mi 48109-0720 (USA) (Accepted 10 March 1992) Key words: /3-Endorphin: Nucleus tractus solitarius; Pro-opiomelanocortin mRNA Evidence from a number of sources indicates that the major site of pro-opiomelanocortin (POMC)-producing cells in the CNS is the arcuate nucleus of the hypothalamus. Using immunocytochemical techniques, a second, smaller group of POMC cells has been detected in the nucleus tractus solitarius (NTS) area of the caudal medulla. However, POMC mRNA has never been reported in the NTS even though it has been found in other extrahypothalamic brain regions. Thus, there is some uncertainty as to whether POMC peptides are actually synthesized de nero in the NTS. in the present study, we used biochemical and anatomical techniques to examine whether POMC mRNA is localized in the NTS. Using in situ hybridization, cells containing POMC mRNA were found in the caudal portion of the NTS. The nucleic acid distribution correlated well with the anatomical distribution of 16k POMC peptide immunoreactivity as determined by immunocytochemistry. Northern analysis revealed that the apparent size of POMC mRNA in the NTS was similar to that found in the arcuate nucleus or the pituitary gland. Results of RNase protection assays using a POMC riboprobe complementary to the 5' end of exon 3 suggested that POMC mRNA in the NTS and arcuate nucleus are identical in this region of the message at least. We also calculated POMC peptide product to mRNA ratios in different tissues and found that NTS cells appear to produce less peptide per mRNA molecule than those in the arcuate nucleus or pituitary gland. Taken togelher, these data provide the strongest evidence to date that POMC is synthesized de nero in the NTS and offer support for a role for POMC.derived opioid and non-opioid peptides in autonomic functions in the caudal medulla. INTRODUCTION The potent opioid peptide /J.endorphin (fiE) is derived from the precursor protein pro.opiomelano- cortin (POMC). While there is abundant evidence that /3E may be a physiologically important peptide trans- mitter involved in diverse behaviors such as stress, pain, feeding, and autonomic responses (for review, see refs. 2 and 13), it has been difficult to define specific POMC pathways and to associate them with particular functions. Part of this difficulty relates to confusion about the precise localization of POMC somata and the anatomical distribution of their axonal projections. The major POMC cell group is found in the arcuate region of the mediobasal hypothulamus .~,4,14,2(,.2,),.~(,-.~s. Axonal fibers from this cell group project to a number of brain areas including the septum, amygdala, various hypothalamic and thalamic nuclei and the midbrain. The discovery that POMC mRNA was also found in the arcuate nucleus ~(' suggested that these cells intrin- sically synthesized POMC peptides (as opposed, for example, to taking them up from the extracellular space). This hypothesis was confirmed by two sets of data: first, Liotta et al. demonstrated by pulse labelling techniques that cells in the arcuate could synthesize and process POMC peptides de nero; and second, using immunocytochemical (ICC) and in situ hybridiza- tion (ISH) techniques, cells in the arcuate were shown to contain both POMC mRNA and peptide product ii. At the present time, it is not clear whether other brain sites also contain cells which synthesize POMC peptide products de novo. Using ICC, several groups reported the existence of a second cluster of POMC cells in the nucleus tractus solitarius (NTS) area of the Correspondence: D.M. Bronstein. Present address: LMIN, NIEHS, NIH, P.O. Box 12233, Mail Drop 14-06, Research Triangle Park, NC 27709, USA. * Present address: Anatomisches lnstitut, Johannes Gutenbe~g Universitat, Saarstrasse 21, D-6500, Mainz, Germany.