Margaret S. Kreider

Reduction of feeding behavior by the serotonin uptake inhibitor sertraline

Administration of the selective serotonin (5-HT) uptake inhibitor sertraline produced a dose-dependent reduction of food intake in rats. Doses of sertraline of 10 mg/kg or greater reduced the intake of solid pellets significantly (P<0.01) during the 1st hour of a 4-h feeding test in rats deprived of food and water for 24 h. Food intake during the remaining 3 h and water intake during the feeding test was unaffected by sertraline. Sertraline (2–18 mg/kg IP) also reduced milk consumption in food-deprived rats. Pretreatment with the nonselective 5-HT antagonists metergoline (2 mg/kg IP) or methysergide (3.3 mg/kg IP) blocked sertralines inhibition of dry food intake, whereas pretreatment with the selective 5-HT2 receptor antagonist ketanserin (3.3 mg/kg IP) or the peripheral 5-HT2 antagonist xylamidine (2.5 mg/kg IP) failed to block sertralines anorexic effect. The feeding-suppressant effect of 10 mg/kg sertraline was prevented following the destruction of central 5-HT neurons by the 5-HT neurotoxic agent, 5,7-dihydroxytryptamine (200 μg ICV). This result is consistent with sertralines anorexic effect depending on intact 5-HT neurotransmission. Therefore, sertraline appears to reduce feeding by enhancing the action of endogenous serotonin at central synapses mediated by 5-HT1 rather than 5-HT2 receptors.

Ultrastructural localization of thyrotropin-releasing hormone immunoreactivity in the dorsal vagal complex in rat

Thyrotropin-releasing hormone-like immunoreactivity (TRH-LI) was localized at the ultrastructural level in the dorsal vagal complex (DVC: dorsal motor nucleus of the vagus (DMV) and the nucleus of the solitary tract (NST] in rat. TRH-LI was concentrated in large granular vesicles in axons, presynaptic terminals, and non-synaptic axon varicosities. TRH-LI presynaptic terminals established both asymmetric and symmetric synaptic contacts with dendrites. These observations are consistent with recently described direct inhibitory and facilitatory effects of TRH on the electrical activity of neurons in the DVC.

Effects of intrabulbar injections of 6-hydroxydopamine on ethyl acetate odor detection in castrate and non-castrate male rats

The function of norepinephrine-containing neurons which project to the olfactory bulb is poorly understood. Although there has been suggestion that norepinephrine (NE) may modulate general olfactory sensitivity by attenuating the inhibitory feedback of granule cells upon mitral and tufted cells, behavioral indices of olfactory sensitivity have not been measured in animals with depletions of bulbar NE. The present experiment used computerized olfactometry and signal detection methodology to assess the odor detection performance of castrate and non-castrate male rats to a range of perithreshold concentrations of ethyl acetate following 6-hydroxydopamine (6-OHDA) depletion of bulbar NE. Such depletion had no significant influence on odor detection performance at any of the odorant concentrations examined in either castrate or non-castrate animals, as indexed by the non-parametric sensitivity measure SI. This observation implies that general olfactory sensitivity is unaltered by major depletion of intrabulbar NE, but does not preclude the possibility that NE modulates sensitivity to select odorants or odorant mixtures, or alters detection ability under atypical states of arousal.

Immunohistochemical localization of TRH in rat CNS: Comparison with RIA studies ☆

The localization of thyrotropin releasing hormone (TRH) in rat brain determined by use of avidin-biotin immunoperoxidase histochemistry was compared with the distribution and quantitation by radioimmunoassay (RIA). Male Sprague-Dawley rats received intracisternal injections of 100 micrograms of colchicine or saline and were sacrificed 24 hours later. Brains were either perfused with lysine-periodate fixative and processed for TRH immunohistochemistry or were dissected into 9 brain regions for TRH RIA. In colchicine pretreated rats. TRH immunoreactive perikarya were observed only in nuclei of the hypothalamus and brain stem. No cell body staining was observable in non-colchicine treated rats. With the exception of the olfactory bulb, brain regions exhibiting dense TRH staining contained high concentrations of TRH as measured by RIA. Colchicine pretreatment did not alter the concentration of TRH in most brain regions, however, there was a significant increase in brain stem TRH content 24 hours following colchicine administration. These findings indicate that immunohistochemical localization of TRH corresponds well with endogenous concentrations of TRH determined by RIA.

TRH immunoreactivity in rat hypothalamus and brain: assessment by gel filtration and thin-layer chromatography

Thyrotropin-releasing hormone (TRH) has a number of central nervous system neuropharmacological and behavioral actions 4. Interest in TRH as a possible endogenous neuromodulator was increased by the finding of TRH immunoreactivity (IR-TRH) throughout the central nervous system 2,z,5. Extrahypothalamic and hypothalamic IR-TRH and synthetic TRH have similar physiochemical and biological properties in a number of systems, including gel filtration, electrophoresis and bioassay. These findings provided indirect but strong support that hypothalamic and extrahypothalamic IR-TRH was indeed TRH. Recently, Youngblood et al, reported studies suggesting that IR-TRH differed from synthetic TRH 6. Specifically, IR-TRH from brain extracts did not co-migrate with synthetic TRH on thin-layer chromatography and could not be recovered by affinity chromatography of brain extracts. These data were the first suggesting that extrahypothalamic IR-TRH differed from synthetic TRH. We have repeated these studies, but were unable to discern differences in extrahypothalamic and hypothalamic IR-TRH and synthetic TRH. Some suggestions are offered for the differences between our findings and those of Youngblood et al. 6. Male Sprague-Dawley rats (180-200 g) were decapitated, the brains removed, and extracts of whole brain, extrahypothalamic brain, frontal cortex or hypothalamus were prepared in two ways. Whole brains were each homogenized in 2 M acetic acid (5 ml/g tissue), as described by Youngblood et al. 6. After centrifugation at 2000 r.p.m. for 10 min, the supernatants were extracted with an equal volume of diethyl ether, frozen, and thawed to break the interphase. The aqueous layer was lyophilized. The dried residue was extracted with 15 ml ethanol, centrifuged at 2000 r.p.m, and the supernatants dried overnight at 60 °C in an air stream. Hypothalami, frontal cortices, extrahypothalamic brains, and whole brains also were each homogenized in 2.0 ml 0.15 M NaCI, 0.01 M NaH2PO4, pH 7.5, and extracted with 10 ml methanol. The homogenates were centrifuged at 2000 r.p.m, for 10 min and the supernatants dried

The olfactory bulb is rich in TRH immunoreactivity

We report that the rat olfactory bulb is rich in thyrotropin releasing hormone (TRH) immunoreactivity. TRH content was determined according to the radioimmunoassay method of Bassiri and Utiger. The concentration (mean +/- SEM., n = 10) of TRH in olfactory bulb (60 +/- 10 pg/mg wet weight) was 23% of the concentration in the hypothalamus, and was at least twice that of other brain regions examined. The 2 olfactory bulbs (mean wet weight 65 mg/2 bulbs) contained 3.9 +/- 0.3 ng TRH. The TRH immunoreactivity could be separated into high and low molecular weight forms. The low molecular weight form co-chromatographed with authentic TRH (mol. wt. 362) on gel filtration and thin layer adsorption chromatography and caused the release of thyrotropin from pituitary tissue incubated in vitro. Since the neuronal organization and functions of the olfactory bulb are well described, studies of the localization and metabolism of TRH in this region may help to clarify the role of this tripeptide in the central nervous system.

Systemic administration of kainic acid produces elevations in TRH in rat central nervous system

Several studies have suggested that the concentration of thyrotropin releasing hormone (TRH) in the central nervous system (CNS) is influenced by the level of CNS activation. Hibernation in the ground squirrel and estivation in the lungfish result in region-specific decreases in TRH concentrations. Repeated electroconvulsive shock (ECS) and amygdaloid kindling have been shown to result in elevations of TRH in limbic brain regions. In the present study, limbic seizures induced by systemic administration of kainic acid resulted in substantial increases in the TRH content of posterior cortex and of dorsal and ventral hippocampus, and in moderate elevations in anterior cortex, amygdala/piriform cortex and corpus striatum. Maximal elevations in TRH were observed 2-4 days after kainic acid administration, and by 14 days TRH levels were similar to control values, with the exception of the dorsal hippocampus, which exhibited more prolonged elevations in TRH levels. Prior exposure to limbic seizure activity attenuated the magnitude of TRH elevation in response to a second administration of kainic acid in the posterior cortex but in no other region. These results indicate that seizure-related processes or events influence TRH systems in the CNS. Neuronal populations involved in limbic seizure induced damage may be involved in the modulation of posterior cortical TRH levels.

TRH and TRH Receptors in the Spinal Cord

Over the past 12 years, substantial progress has been made in delineating the localization of TRH and TRH receptors in spinal cord. High concentrations of both the peptide and its receptor have been observed in the ventral horn in the region of the motoneurons and in the dorsal horn in the substantia gelatinosa. As noted, pharmacological effects of TRH administration on various parameters of spinal cord function have been reported in a number of studies. To date, however, substantial questions remain regarding the physiological role of TRH in the spinal cord. Nevertheless, it is hoped that the extensive information that has been obtained on localization of TRH and TRH receptors in spinal cord will provide a basis for answering these complex questions.

Pathways of TRH degradation in rat brain

Abstract We have investigated the TRH degradative enzymes in brain by examining the pattern of metabolites formed in vitro . The homogenate and three subcellular fractions of rat brain were separately incubated for 1, 5, and 15 minutes with 3H-TRH (pyro-glu-his-3H-proNH2) at 37°C. TRH and its metabolites were separated on silica gel thin-layer chromatography plates. The crude homogenate and subcellular fractions each produced a characteristic pattern of metabolism. The homogenate metabolized TRH to TRH-OH, proline, and prolineamide. With the P1 fraction, prolineamide and proline were the major metabolites. In both the homogenate and P1 fraction incubations, histidyl-prolineamide appeared as a minor component. The P2 fraction produced prolineamide and histidyl-prolineamide as the major metabolites while the cytosol metabolized TRH primarily to TRH-OH. Proline was formed during incubation with both cytosol and P2 fractions. The TRH deamidase is found in the soluble fraction of brain tissue homogenate while the pyroglutamate aminopeptidase and the prolineamide cleaving enzyme are associated with particulate fractions. Histidyl-prolineamide is further degraded in the homogenate and P1 fractions by a secondary metabolic pathway. Proline salvaging enzymes are present in all subcellular fractions of rat brain.

Destruction of the nucleus raphe obscurus and potentiation of serotonin-mediated behaviors following administration of the neurotoxin 3-acetylpyridine

Systemic administration of the neurotoxin 3-acetylpyridine (3-AP) to rats produced spontaneous episodes of spasmodic movement involving the trunk and limbs including torticollis, contortions of the trunk and rigid extension of the limbs. Because the neurotransmitter serotonin (5-HT) has been implicated in various human involuntary movement disorders, the functional and anatomical integrity of the 5-HT system in rats treated with 3-AP were examined. 5-HT-containing neurons in the brain stem were studied using immunohistochemical labeling with antiserum to 5-HT. Cells in the nucleus raphe obscurus were found to be altered following 3-AP treatment as shown by a decrease in 5-HT immunoreactivity as compared to control rats. No changes in 5-HT immunoreactivity were observed in any other region containing 5-HT cell bodies. Behaviorally, rats treated with 3-AP were 2.5-fold more sensitive to the ability of the 5-HT1A agonist 8-OH-2-(di-n-propylamino)tetralin (8-OH-DPAT; 0.33-3.3 mg/kg) to produce the 5-HT syndrome. Similarly, 3-AP-treated rats were 2-fold more sensitive to the selective 5-HT2 agonist 1-(2,5-dimethoxy-4-bromophenyl)-2-aminopropane (DOB; 0-1.0 mg/kg) at producing the head shake response. Although these behaviors associated with brain stem 5-HT receptors were potentiated by 3-AP, the hypothermic effect of 8-OH-DPAT which involves ascending mesencephalic 5-HT neurons was unchanged following 3-AP treatment. Treatment with 3-AP did not produce significant alterations of 5-HT or 5-hydroxyindoleacetic acid (5-HIAA) content in any brain region studied. Quantitative autoradiographic analysis of the density of 5-HT1A receptors labeled with [3H]8-OH-DPAT revealed that these sites were unchanged in regions of the brain (frontal cortex, hippocampus and brain stem) and in the spinal cord. Similarly, few changes in the density of 5-HT2 receptors measured with [3H]ketanserin were observed in various brain regions. These results suggest that neurons from the nucleus raphe obscurus are involved in the elicitation of 5-HT-mediated behavioral responses by 5-HT1A and 5-HT2 receptor agonists that are though to be mediated through brain stem and spinal cord mechanisms. In addition, because of the close neuroanatomical relationship of the nucleus raphe obscurus with various brain regions known to be involved in motor control, the destruction of this region by 3-AP may contribute to the spasmodic motor behaviors observed following 3-AP treatment.