The activation of the HPA axis is the endocrine measure of stress responsiveness that is initiated by corticotropin-releasing hormone (CRH). CRH exerts its effects via CRHR1 and CRH-R2 receptors coupled to the cAMP signaling system and this process involves transcription factor cAMP-responsive element-binding protein (CREB).This study investigated the role of CRH and the possible involvement of CREB in gene regulation of CRH receptor, under basal conditions and after stress application in the pituitary. We used wild type (wt +/+) controls and CRH knock-out (CRH-KO -/-) male mice. Using CRH-deficient mice, we were able to investigate the consequences of the lack of the CRH on the expression of CRH receptors and transcriptional regulation mediated by CREB. We estimated the effect of acute (IMO 1×) and repeated (IMO 7×) restraint stressors lasting 30 and 120 min on the expression of mRNA CREB, CRH-R1, and CRH-R2 by qPCR. We found very significant difference in the expression of these peptides under the effect of single and repeated stress in control and CRH-KO mice. Our results indicate that both CRH receptors and CREB might be involved in the regulation of stress response in the pituitary of mice. We propose that regulation of the stress response may be better understood if more were known about the mechanisms of CRH receptor signal transduction and involvement of CREB system.
- MeSH
- akutní nemoc MeSH
- hormon uvolňující kortikotropin biosyntéza nedostatek MeSH
- hypofýza metabolismus MeSH
- myši inbrední C57BL MeSH
- myši knockoutované MeSH
- myši MeSH
- protein vázající cAMP responzivní element biosyntéza MeSH
- psychický stres metabolismus psychologie MeSH
- receptory hormonu uvolňujícího kortikotropin biosyntéza MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Brain acetylcholinesterase (AChE) variant AChERexpression increases with acute stress, and this persists for an extended period, although the timing, strain and laterality differences, have not been explored previously. Acute stress transiently increases acetylcholine release, which in turn may increase activity of cholinesterases. Also the AChE gene contains a glucocorticoid response element (GRE), and stress-inducible AChE transcription and activity changes are linked to increased glucocorticoid levels. Corticotropin-releasing hormone knockout (CRH-KO) mice have basal glucocorticoid levels similar to wild type (WT) mice, but much lower levels during stress. Hence we hypothesized that CRH is important for the cholinesterase stress responses, including butyrylcholinesterase (BChE). We used immobilization stress, acute (30 or 120 min) and repeated (120 min daily × 7) in 48 male mice (24 WT and 24 CRH-KO) and determined AChER, AChE and BChE mRNA expression and AChE and BChE activities in left and right brain areas (as cholinergic signaling shows laterality). Immobilization decreased BChE mRNA expression (right amygdala, to 0.5, 0.3 and 0.4, × control respectively) and AChERmRNA expression (to 0.5, 0.4 and 0.4, × control respectively). AChE mRNA expression increased (1.3, 1.4 and 1.8-fold, respectively) in the left striatum (Str). The AChE activity increased in left Str (after 30 min, 1.2-fold), decreased in right parietal cortex with repeated stress (to 0.5 × control). BChE activity decreased after 30 min in the right CA3 region (to 0.4 × control) but increased (3.8-fold) after 120 min in the left CA3 region. The pattern of changes in CRH-KO differed from that in WT mice.
- MeSH
- acetylcholinesterasa metabolismus MeSH
- butyrylcholinesterasa metabolismus MeSH
- funkční lateralita fyziologie MeSH
- fyzické omezení MeSH
- fyziologický stres fyziologie MeSH
- hormon uvolňující kortikotropin genetika metabolismus MeSH
- mozek metabolismus MeSH
- myši knockoutované MeSH
- myši MeSH
- psychický stres metabolismus MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Glucocorticoids act via glucocorticoid receptors (GR), typically localized in the cytosol (cGR). Rapid action is probably mediated via membrane receptors (mGR). In corticotropin-releasing hormone knockouts (CRH-KO), basal plasma glucocorticoid levels do differ from wild type levels (WT), but are approximately ten times lower during exposure to immobilization stress (IMMO) in comparison to WT. We tested the following hypotheses: (1) the mice lung tissue GR basal numbers would not be changed in CRH-KO (because of similar glucocorticoid levels), (2) the number of GR would be changed in WT but not in KO during short (30, 90, and 120 min) IMMO (because of higher increase of glucocorticoid levels in WT). The basal levels of cGR were not changed in CRH-KO (compared to WT), while mGR were significantly lower (62 %) in CRH-KO. In WT, there was the only decrease (to 32 %) in cGR after 120 min when we also found an increase in mGR in WT (to 201 %). In CRH-KO, IMMO caused gradual decrease in cGR (to 52 % after 30 min, to 46 % after 90 min, and to 32 % after 120 min). In CRH-KO, the only increase in mGR appeared already at 30 min of IMMO. These data suggest, on the contrary to our hypotheses, that CRH-KO are more susceptible to GR changes in early phases of stress.
- MeSH
- buněčná membrána metabolismus MeSH
- cytosol metabolismus MeSH
- dexamethason metabolismus MeSH
- fyziologický stres * MeSH
- glukokortikoidy metabolismus MeSH
- hormon uvolňující kortikotropin metabolismus MeSH
- imobilizace * MeSH
- kinetika MeSH
- myši knockoutované MeSH
- myši MeSH
- plíce metabolismus MeSH
- receptory glukokortikoidů metabolismus MeSH
- vazebná místa MeSH
- western blotting MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Although c-Fos plays a key role in intracellular signalling, the disruption of the c-fos gene has only minor consequences on the central nervous system (CNS) function. As muscarinic receptors (MR) play important roles in many CNS functions (attention, arousal, and cognition), the c-fos knock-out might be compensated through MR changes. The aim of this study was to evaluate changes in the M1-M5 MR mRNA in selected CNS areas: frontal, parietal, temporal and occipital cortex, striatum, hippocampus, hypothalamus and cerebellum (FC, PC, TC, OC, stria, hip, hypo, and crbl, respectively). Knocking out the c-fos gene changed the expression of MR in FC (reduced M1R, M4R and M5R expression), TC (increased M4R expression), OC (decreased M2R and M3R expression) and hippocampus (reduced M3R expression). Moreover, gender differences were observed in WT mice: increased expression of all M1-M5R in the FC in males and M1-M4R in the striatum in females. A detailed analysis of MR transcripts showed pre-existing correlations in the amount of MR-mRNA between specific regions. WT mice showed three major types of cortico-cortical correlations: fronto-occipital, temporo-parietal and parieto-occipital. The cortico-subcortical correlations involved associations between the FC, PC, TC and striatum. In KO mice, a substantial rearrangement of the correlation pattern was observed: only a temporo-parietal correlation and correlations between the FC and striatum remained, and a new correlation between the hypothalamus and cerebellum appeared. Thus, in addition to the previously described dopamine receptor restructuring, the restructuring of MR mRNA correlations reveals an additional mechanism for adaptation to the c-fos gene knockout.
- MeSH
- genetická transkripce MeSH
- geny fos * genetika MeSH
- kvantitativní polymerázová řetězová reakce MeSH
- messenger RNA MeSH
- mozek metabolismus MeSH
- myši inbrední C57BL MeSH
- myši knockoutované MeSH
- myši MeSH
- pohlavní dimorfismus MeSH
- receptory muskarinové biosyntéza genetika MeSH
- regulace genové exprese genetika MeSH
- transkriptom MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Although c-Fos protein is one of the principal molecules in intracellular signaling, c-fos gene disruption is associated with alterations in neuronal functions that do not correspond to its importance in function. The aim of the study was to evaluate the changes of dopaminergic system together with acetylcholinesterase (AChE) in c-fos disruption (KO). KO male mice showed an increase in D₁-like receptor (279% of WT) and D₂-like receptor (345% of WT) binding sites in the cortex. On the gene expression level (assessed by real-time PCR), lower quantities of D₁R-mRNA (0.64) and D₅R-mRNA (0.6) were found in females when compared to males in the frontal cortex, higher D₂R-mRNA in the parietal (1.43) and temporal (2.64) cortex and lower AChE-mRNA (0.67). On the contrary, female striatum contained higher level of D₂R-mRNA (1.62) and AChE-mRNA (1.57) but lower level of D₃R-mRNA (0.73). Hypothalamic D₁R-mRNA, D₂R-mRNA and D₄R-mRNA were higher in females (1.38, 1.63, and 1.68, respectively). Disruption of c-fos increased selectively D₅R-mRNA (1.31) in male parietal cortex, D₂R-mRNA (1.72) in male temporal cortex, and cerebellar D₂R-mRNA in both males (1.43) and females (1.42), respectively. In females, we found rather decrease in DR-mRNA. Multiple correlations in mRNA quantities (in WT mice) were found, which changed considerably upon c-fos KO. Main interactions in WT were inter-regional, CNS of KO underwent an extensive restructuring comprising intraregional interactions in the frontal cortex, hypothalamus, and cerebellum. These changes in DR (between others) could be considered as one of the adaptive mechanisms in c-fos KO mice.
- MeSH
- acetylcholinesterasa genetika metabolismus MeSH
- antagonisté dopaminu farmakokinetika MeSH
- benzazepiny farmakokinetika MeSH
- mapování mozku MeSH
- messenger RNA metabolismus MeSH
- mozek účinky léků metabolismus MeSH
- myši inbrední C57BL MeSH
- myši knockoutované MeSH
- myši MeSH
- protoonkogenní proteiny c-fos nedostatek MeSH
- receptory dopaminové genetika metabolismus MeSH
- regulace genové exprese účinky léků genetika MeSH
- sexuální faktory MeSH
- spiperon farmakokinetika MeSH
- tritium farmakokinetika MeSH
- vazba proteinů účinky léků genetika MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
We investigated the role of beta3-adrenoceptors (AR) in cold stress (1 or 7 days in cold) in animals lacking main cardioinhibitive receptors-M2 muscarinic receptors (M(2)KO). There was no change in receptor number in the right ventricles. In the left ventricles, there was decrease in binding to all cardiostimulative receptors (beta1-, and beta2-AR) and increase in cardiodepressive receptors (beta3-AR) in unstressed KO in comparison to WT. The cold stress in WT animals resulted in decrease in binding to beta1- and beta2-AR (to 37%/35% after 1 day in cold and to 27%/28% after 7 days in cold) while beta3-AR were increased (to 216% of control) when 7 days cold was applied. MR were reduced to 46% and 58%, respectively. Gene expression of M2 MR in WT was not changed due to stress, while M3 was changed. The reaction of beta1- and beta2-AR (binding) to cold was similar in KO and WT animals, and beta3-AR in stressed KO animals did not change. Adenylyl cyclase activity was affected by beta3-agonist CL316243 in cold stressed WT animals but CL316243 had almost no effects on adenylyl cyclase activity in stressed KO. Nitric oxide activity (NOS) was not affected by BRL37344 (beta3-agonist) both in WT and KO animals. Similarly, the stress had no effects on NOS activity in WT animals and in KO animals. We conclude that the function of M2 MR is substituted by beta3-AR and that these effects are mediated via adenylyl cyclase rather than NOS.
- MeSH
- adenylátcyklasy metabolismus MeSH
- beta-3-adrenergní receptory metabolismus MeSH
- fyziologická adaptace genetika MeSH
- fyziologický stres genetika MeSH
- katecholaminy biosyntéza MeSH
- myši MeSH
- nízká teplota MeSH
- receptor muskarinový M2 nedostatek genetika metabolismus MeSH
- regulace genové exprese MeSH
- srdce patofyziologie MeSH
- srdeční komory enzymologie patologie patofyziologie MeSH
- synthasa oxidu dusnatého metabolismus MeSH
- vazba proteinů MeSH
- vazebná místa MeSH
- zvířata MeSH
- Check Tag
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
We tested the hypothesis that single and repeated immobilization stress affect densities of alpha(1)-adrenoceptor (alpha(1)-AR) and beta-AR subtypes, muscarinic receptors (MR), adenylyl cyclase activity (AC) and phospholipase C activity (PLC) in lungs of male and female wild type (WT) and corticotropin-releasing hormone gene (CRH-knockout (KO)) disrupted mice. We found sex differences in the basal levels of alpha(1)-AR subtypes (females had 2-3 times higher density of receptors than males) and MR (males had twice the density found in females). In marked contrast, beta-AR subtype densities did not differ between sexes. CRH gene disruption decreased all three studied receptors in intact mice (to 20-50% of WT) in both sexes (except beta(1)-AR in females). Stress induced sexually dimorphic responses, while all alpha(1)-AR subtypes decreased in females (to 30% of control approximately), only alpha(1A)-AR level diminished (about 50%) in males. beta(1)-AR decreased in males (to about 40%) but remained stable in females. beta(2)-AR diminished in females (to about 20-60%) and also in males (to about 30-60%). MR decreased in both sexes (approximately to 50%). AC activity diminished in males (to < 50%) while PLC activity was not changed. In CRH-KO mice, the stress response was severely diminished. Paradoxically, the receptor response to stress was less affected by CRH-KO in males than in females. AC activity did not change in CRH-KO mice. In conclusion, in mice the stress reaction is sexually dimorphic and an intact hypothalamo-pituitary-adrenocortical system is required for the normal reaction of pulmonary adrenergic and MR to stress.
- MeSH
- alfa-1-adrenergní receptory metabolismus MeSH
- beta-adrenergní receptory metabolismus MeSH
- fyzické omezení MeSH
- fyziologický stres fyziologie MeSH
- hormon uvolňující kortikotropin genetika metabolismus MeSH
- myši knockoutované MeSH
- myši MeSH
- plíce metabolismus MeSH
- pohlavní dimorfismus MeSH
- receptory muskarinové metabolismus MeSH
- sexuální faktory MeSH
- systém hypofýza - nadledviny metabolismus MeSH
- systém hypotalamus-hypofýza metabolismus MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
