Effect of dexfenfluramine on the transcriptional activation of CRF and its type 1 receptor within the paraventricular nucleus of the rat hypothalamus.
1. The present study investigated the effect of intraperitoneal (i.p.) administration of the indirect 5-hydroxytryptamine (5-HT) receptor agonist, dexfenfluramine, on the transcriptional activity of corticotropin-releasing factor (CRF) and its type 1 receptor in the brains of conscious male Sprague-Dawley rats via in situ hybridization histochemistry (ISHH) using both intronic and exonic probe technology. 2. The immediate early gene (IEG) c-fos mRNA was also used as index of cellular activity, whereas localization between CRF-immunoreactive (ir) perikarya and the IEG was accomplished to determine the site of CRF neuronal activation in the brain of dexfenfluramine-treated rats. 3. Thirty minutes, 1, 3, and 6 h after a single injection of either dexfenfluramine (10 mg kg-1) or the vehicle solution, adult male rats (230-260 g) were deeply anaesthetized and rapidly perfused with a 4% paraformaldehyde-borax solution (PF). The brains were removed from the skull, postfixed, and placed in a solution of 4% PF-10% sucrose overnight at 4 degrees C. Frozen brains were mounted on a microtome and cut from the olfactory bulb to the medulla in 30-microns coronal sections. 4. Dexfenfluramine induced a general neuronal activation as indicated by the strong signal of c-fos mRNA in several structures of the brain, including the parietal cortex, caudate putamen, circumventricular organs, medial preoptic area, bed nucleus of the stria terminalis, choroid plexus, choroidal fissure, supraoptic nucleus, paraventricular nucleus of the hypothalamus (PVN), paraventricular nucleus of the thalamus, central nucleus of the amygdala, dorsomedial nucleus of the hypothalamus, laterodorsal tegmental nucleus, locus coeruleus, and several subdivisions of the dorsal vagal complex. In most of these structures, the signal was maximal at 30 min, still strong and positive at 60 min, largely decreased at 3 h, and had completely disappeared 6 h after injection. 5. In the parvocellular division of the PVN, the large majority of CRF-ir perikarya displayed a positive signal for the mRNA encoding c-fos, indicating a profound CRFergic activation within this neuroendocrine nucleus after dexfenfluramine administration. 6. Colocalization between CRF-ir neurones and c-fos positive cells was not detected in any other regions. This selective activation of PVN CRF neurones was also confirmed by the presence of CRF primary transcript; 30 min after i.p. injection of the indirect 5-HT agonist, a positive signal for CRF hnRNA was observed, specifically in the parvocellular PVN. 7. Transcription of the gene encoding the type 1 receptor for CRF was highly stimulated in the PVN following 5-HT activation. Although this hypothalamic nucleus exhibited a barely detectable signal under basal conditions, dexfenfluramine induced a strong signal of CRF1 receptor mRNA in the parvocellular PVN. Interestingly, CRF-ir neurones displayed a positive signal for the mRNA encoding the CRF1 receptor, 3 and 6 h after systemic treatment with dexfenfluramine. 8. These results indicate that although dexfenfluramine can generate a wide neuronal activation throughout the brain, this 5-HT agonist triggers the activity of CRF neurones selectively in the parvocellular division of the PVN, a mechanism possibly related to the activity of hypothalamic-pituitary-adrenal axis. Induction of CRF1 receptor mRNA in CRF cells of the PVN indicates that neuroendocrine CRF neurones can be targeted by CNS CRF under 5-HT stimulation.
Laflamme
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N
Laboratory of Molecular Endocrinology, CHUL Research Center, Laurier, Québec, Canada.
Bovetto
S
S
Richard
D
D
Rivest
S
S
D000818:Animals; D003346:Corticotropin-Releasing Hormone / Q000235:genetics*; D005277:Fenfluramine / Q000494:pharmacology*; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D008297:Male; D010286:Paraventricular Hypothalamic Nucleus / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D016760:Proto-Oncogene Proteins c-fos / Q000096:biosynthesis / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D018019:Receptors, Corticotropin-Releasing Hormone / Q000096:biosynthesis / Q000235:genetics*; D017366:Serotonin Receptor Agonists / Q000494:pharmacology*; D013997:Time Factors; D014158:Transcription, Genetic / Q000187:drug effects*
D016760:Proto-Oncogene Proteins c-fos; D012333:RNA, Messenger; D018019:Receptors, Corticotropin-Releasing Hormone; D017366:Serotonin Receptor Agonists; D005277:Fenfluramine; D003346:Corticotropin-Releasing Hormone