No abstract

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 novo 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.(ABSTRACT TRUNCATED AT 250 WORDS)

In previous studies to determine whether chronic opiate administration might negatively feedback upon endogenous opioid systems in the CNS, investigators found no changes in steady-state concentrations of opioid peptides following morphine pelleting. However, since only steady-state levels were measured, it was still not clear whether morphine treatment altered the release and/or biosynthesis of opioid-containing neurons. The goal of the present study was to assess the effects of chronic morphine pelleting on the dynamics of beta-endorphin (beta E) biosynthesis in rats. Hence, at several times during a 7-day morphine treatment, concentrations of total beta E-immunoreactivity (-ir), as well as chromatographically sieved forms of beta E, were determined by RIA, and mRNA levels of pro-opiomelanocortin (POMC) were measured by a solution phase protection assay using a mouse or rat POMC 32P-labelled riboprobe. Concentrations of total beta E-ir or different forms of beta E-ir peptides (i.e. beta-lipotropin, beta E1-31, or beta E1-27/beta E1-26) in the hypothalamus or midbrain following either 1 or 7 days of treatment were similar in morphine- and placebo-pelleted animals. However, a significant increase in total hypothalamic beta E-ir was observed following 3 days of morphine pelleting; chromatographic analyses indicated that this was primarily due to a selective increase in the opiate inactive forms of beta E, i.e. beta E1-27/beta E1-26. After 7 days of pelleting, morphine-treated animals tended to have lower POMC mRNA levels than those of placebo controls (20 to 50% decrease in different studies). The accumulation of hypothalamic beta E-ir at 3 days, and the apparent decline in POMC mRNA levels at 7 days, lend support to the hypothesis that morphine negatively feeds back upon POMC neurons in the brain by inhibiting beta E release and biosynthesis.

The distribution of preprodynorphin messenger RNA-containing perikarya in the central nervous system of the rat was determined with in situ hybridization histochemistry using a 35S-labelled complementary RNA probe. All of the regions of the central nervous system reported by other investigators to contain perikarya that synthesize prodynorphin-derived peptides, except the pedunculopontine tegmental nucleus, the accessory trigeminal nucleus, and the ventral nucleus of the trapezoid body, also contained perikarya that synthesize preprodynorphin messenger RNA. However, the olfactory bulb, the anterior olfactory nucleus, the islands of Calleja, the CA1-CA3 fields of the hippocampus, the septohippocampal nucleus, the diagonal band of Broca, the basal and cortical amygdaloid nuclei, the entopeduncular nucleus, the subthalamic nucleus, the superior colliculus, the Edinger-Westphal nucleus, the dentate nucleus, the raphes linearis and pontis, the dorsal cochlear nucleus, the medial vestibular nucleus, the inferior olive, and the dorsal motor nucleus of the vagus nerve also contained preprodynorphin messenger RNA-synthesizing perikarya. These observations suggest that prodynorphin-derived peptides have a much more pervasive role in central nervous system function than previously suspected. However, before the physiological significance of these observations can be judged, it will be necessary to determine whether all of the novel sites of preprodynorphin messenger RNA synthesis are sites of prohormone synthesis and conventional processing.

Maturational changes in visceral nociception were measured in developing or adult rats challenged with hypertonic saline or acetylcholine. Chemically induced abdominal constrictions were absent in rats younger than 7 days of age, regardless of the dose of hypertonic saline or acetylcholine used. Age-related increases in the percent of animals responding, the number of abdominal constrictions emitted per responder, and total response duration occurred in animals 10-20 days of age, until adult-like patterns of responding were attained at the time of weaning. Additional changes in the percent animals responding, as well as in the frequency and total duration of abdominal constriction responses, were also seen in postweanling, but not in preweanling, animals. Five-day-old animals did emit audible vocalizations to the intraperitoneal insertion of a hypodermic needle, however, at a time when these animals failed to show observable responses to the noxious visceral stimuli. Hence, mechanisms mediating pain associated with intraperitoneal needle insertion may be functional during the first postnatal week, at a time when mechanisms mediating visceral pain appear to be immature. These differences may be caused by the differential maturation of sensory, neural, or motoric mechanisms important for hypodermic needle insertion versus visceral nociception.

While enkephalin and dynorphin peptides have been well characterized in the spinal cord, the cellular localization of beta-endorphin (beta E) and the processing of pro-opiomelanocortin (POMC) to beta E and other non-opioid peptides in the cord have not been extensively investigated. Other investigators have characterized the various beta E forms present in rat spinal cord regions. Previous studies have also suggested that spinal POMC content is entirely derived from supraspinal sources. However, high proportions of beta E precursors present in spinal cord sieving profiles led us to suspect the presence of POMC cell bodies intrinsic to the cord. In this study, we performed thoracic spinal cord lesions on a group of animals and demonstrated the persistence of about one-third of control levels of beta E immunoreactivity (beta E-IR) below the level of the lesions. We also characterized POMC processing in various regions of the spinal cord both before and after lesioning. These data suggested that there may be intrinsic POMC/endorphinergic neuronal systems in the spinal cord.

No abstract

There appear to be two anatomically distinct beta-endorphin (beta E) pathways in the brain, the major one originating in the arcuate nucleus of the hypothalamus and a smaller one in the area of the nucleus tractus solitarius (NTS) of the caudal medulla. Previous studies have shown that these two proopiomelanocortin (POMC) systems may be differentially regulated by chronic morphine treatment, with arcuate cells down-regulated and NTS cells unaffected. In the present experiments, we examined the effects of chronic opiate antagonist treatment on beta E biosynthesis across different CNS regions to assess whether the arcuate POMC system would be regulated in the opposite direction to that seen after opiate agonist treatment and to determine whether different beta E-containing areas might be differentially regulated. Male adult rats were administered naltrexone (NTX) by various routes for 8 days (subcutaneous pellets, osmotic minipumps, or repeated intraperitoneal injections). Brain and spinal cord regions were assayed for total beta E-ir, different molecular weight immunoreactive beta-endorphin (beta E-ir) peptides, and POMC mRNA. Chronic NTX treatment, regardless of the route of administration, reduced total beta E-ir concentrations by 30-40% in diencephalic areas (the arcuate nucleus, the remaining hypothalamus, and the thalamus) and the midbrain, but had no effect on beta E-ir in the NTS or any region of the spinal cord. At the same time, NTX pelleting increased POMC mRNA levels in the arcuate to approximately 140% of control values.(ABSTRACT TRUNCATED AT 250 WORDS)