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Alternative splicing of mRNA is one of the most important mechanisms responsible for an increase of the genomic capacity. Thus the majority of human proteins including G protein-coupled receptors (GPCRs) possess several isoforms as a result of mRNA splicing. The corticotropin-releasing factor (CRF) and its receptors are the most proximal elements of hypothalamic-pituitary-adrenal axis (HPA) - the central machinery of stress response. Moreover, expression of CRF and regulated activity of CRF receptor type 1 (CRF1) can also play an important role in regulation of local stress response in peripheral tissues including skin, gastrointestinal tract or reproductive system. In humans, expression of at least eight variants of CRF1 mRNA (α, β, c, d, e, f, g and h) was detected and alternative splicing was found to be regulated by diverse physiological and pathological factors including: growth conditions, onset of labor, during pregnancy or exposure to ultraviolet irradiation. The pattern of expression of CRF1 isoforms is cell type specific and recently has been linked to observed differences in responsiveness to CRF stimulation. In the proposed model of regulation of CRF-signaling, isoform CRF1α plays a central role. Other isoforms modulate its activity by oligomerization, leading to alteration in receptor trafficking, localization and function. Co-expression of CRF1 isoforms modulates sensitivity of cells to the ligands and influences downstream coupling to G-proteins. The other possible regulatory mechanisms include fast mRNA and/or protein turnover or decoy receptor function of CRF1 isoforms. Taken together, alternative splicing of CRF1 can represent another level of regulation of CRF-mediated stress responses at the central and peripheral levels. Chronic stress or malfunction of the HPA-axis have been linked to numerous human pathologies, suggesting that alternative splicing of CRF1 receptor could represent a promising target for drugs development.
The present study indicates that activation of dopamine D1-like receptors by administration of SKF 38393 leads to dose-dependent (doses: 5, 10 and 20 mg/kg) increases in the expression of cFos proteins in the rat paraventricular nucleus of the hypothalamus (PVN). This effect was abolished by administration of SCH 23390, a dopamine D1-like receptor antagonist (0.5 and 1 mg/kg, given 30 min before SKF 38393 - 10 mg/kg), suggesting that the apparent effect is specific for activation of dopamine D1-like receptors. Expression of cFos after SKF 38393 (10 mg/kg) was observed in some, but not all, CRF-immunoreactive neurons, as well as in small portion of oxytocin- but not vasopressin-immunoreactive neurons (double-immunofluorescence experiments). There were also certain populations of nuclei that showed expression of cFos but did not co-localize with the above markers. We also found that both acute and repeated (once daily for 5 consecutive days) exposure to cocaine (25 mg/kg) attenuated the induction of cFos expression triggered by SKF 38393 when administered 24 hours after single or the last dose of cocaine (25 mg/kg). Attenuation was observed at the same level after single and chronic exposure to cocaine, indicating a rapid functional down-regulation of dopamine D1-like receptors that are resistant to subsequent doses of cocaine. These data provide evidence for the functional role of dopamine D1-like receptors in the PVN and indicate a functional adaptation of dopamine D1-like receptors following a single dose of cocaine without further progression of adaptation or resistance of D1-like receptor-mediated genomic function in the course of repeated cocaine intake.
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