| Corticotropin releasing hormone(CRH) and its family of peptides are very important in regulating the central and peripheral nervous system. After the purification of CRH from ovine hypothalamic extracts by Vale in 1981, a series of additional endogenous CRH receptor ligands have also been found, namely, Urocortin1(UCN1), Urocortin2(UCN2) and Urocortin3(UCN3), these amino acid peptide ligands are very similar with CRH on the chain structure. The canonical role of CRH is to initiate the neurohormonal response to stress, via its release from cell bodies within the hypothalamic paraventricular nucleus, which is initiating the hypothalamic-pituitary-adrenal(HPA) axis. In the brain, CRH is not only secreted by the hypothalamus, the central nervous system in other brain regions also found expression of CRH, such as amygdala, hippocampus and basal ganglia. In recent years, the concept, that CRH and its related endogenous peptides and receptor ligands have non-HPA axis actions to regulate CNS synaptic transmission outside the HPA axis, is just beginning to be recognized and identified is especially noteworthy since this synapse has become a prime focus for a variety of mental diseases and neurological disorders, such as schizophrenia and Alzheimer’s disease.Briefly, mammalian CRH receptors have been differentiated broadly into two major types, Corticotropin-Releasing Hormone Receptor Type I(CRHR1) and CorticotropinReleasing Hormone Receptor Type II(CRHR2); each type exhibiting a 70% sequence homology and also possessing molecular splice variants. CRH receptors CRH receptors have the ability to interact with a variety of subunits of G-protein systems. Thus, CRH receptors can modulate various signaling pathways and kinases. The two receptors of CRH have distinct distributions and binding affinities for each receptor. CRH has a high affinity for the R1 receptor and UCN1 binds with high affinity to CRHR1 and CRHR2. While UCN2 and UCN3 have exclusive affinity for the CRHR2. different CRH receptor. Distinct distributions in the brain and the distinct binding affinities with ligands are key determinants of role of CRH peptides.Hippocampus is the key brain region in cognitive, affective and behavioral functions and is a highly plastic structure that is sensitive to the effects of stress and steroid hormones. Synthesis and release of CRH within the hippocampus itself is now established, and its two receptors CRHR1 and CRHR2 widely distribut in the hippocampus, suggesting that the important role of CRH in regulating hippocampal structure and function. Synapses are specialized structures that mediate information flow between neurons and target cells, and thus are the basis for neuronal system to execute various functions, including learning and memory. Studies have shown that stress can lead to hippocampal synaptic morphology changes further affect hippocampal cognitive, emotional responses, and learning and memory functions, and CRH plays a very important role in this process. But the mechanisms are not yet clarified.Increasing body of evidence indicates that stress during pregnancy is associated with increased incidence of a range of affective and behavioral outcomes, such as depression and anxiety, in the offspring in later life(1-4), and elevated concentration of glucocorticoids(GCs) is the key player in this event. Studies have shown that prenatal excess GC exposure impaired spatial learning and caused an increase in depression-like behavior in adult offspring, accompanied by the elevated level of CRH and CRHR1. But the mechanisms of the abnormal hippocampus function and whether high levels of CRH in hippocampus mediated this procedure through the activated CRHR1 have not yet been reported.Therefore, in the first part of this study, Pregnant rats which were received dexamethasone(DEX) in the last week of pregnancy were used as a model and investigated whether neonatal repeated blockade of CRHR1 by CRHR1 antagonist CP154526 or antalarmin could ameliorated impaired spatial learning and hippocampal synaptic abnormalities in adult offspring. We also explored the mechanisms underlying CRH modulation of synaptic formation in hippocampus in different cultured systems, such as hippocampal slices cultures, isolated hippocampal neurons cultures, neuron-astrocyte co-cultures.Studies confirmed that two types of CRH receptor mediated different effects during stress response process. consistently, CRHR1knockout(KO) mice display decreased anxiety-like behaviors. Although CRHR2 KO mice show increased anxiety-like behaviors, UCN2 and UCN3 may have distinct effects on affective behavior including depression and anxiety. In CRHR1 knockout mice, the absence of CRHR1 prevents the detrimental effects of chronic stress on dendritic arborization of CA3 neurons and spatial memory. However, whether UCN2 and UCN3 are involved in synaptic formation by CRHR2 remains unknown. Therefore, in the second part of the experiment of this project, we use a series of experiments to clarify the effects of UCN2 and UCN3 on synaptic formation in hippocampus.Main results:(I) Regulation of spatial learning and memory and synaptic formation by corticotropin-releasing hormone receptor 1 in hippocampus.(1) Regulation of spatial learning and memory by corticotropin-releasing hormone receptor 1. 1. Prenatal excess GC exposure impaired spatial memory acquisition and memory retention in the Morris water maze test in adult offspring, accompanied by a significant increase of CRH and CRHR1 protein level in the hippocampus. 2. The increased CRH and CRHR1 level and the deficit in spatial learning and memory in adult offspring was reversed by neonatal repeated blockade of CRHR1 by CRHR1 antagonist CP154526 or antalarmin.(2) Regulation of synaptic formation by corticotropin-releasing hormone receptor 1 in hippocampus. 1. Prenatal excess GC exposure reduce the level of synapsin I- and PSD95-labeled terminals in CA1 area of hippocampus in adult offspring, and the hippocampal synaptic abnormalities were attenuated by neonatal repeated blockade of CRHR1 by CRHR1 antagonist CP154526 or antalarmin. 2. CRH suppresses presynaptic and postsynaptic protein expression in hippocampal slices by CRHR1. 3. The effect of CRH on synapsin I- and PSD95-labeled terminals formation in isolated neurons differs with that in hippocampal slices, CRH enhanced the numbers and level of synapsin I- and PSD95-labeled terminals by CRHR1. 4. CRH suppresses synapsin I- and PSD95-labeled terminals in neuron-astrocyte co-cultures by CRHR1. 5. Treatment of hippocampal neurons with the media of CRH-treated glias led to an decrease in synapsin I- and PSD95-labeled terminals. 6. CRH treatment led to a reduction of CXCL5/LIX production in glial cells via CRHR1. Administration of CXCL5/LIX prevented suppression of presynaptic terminals in neurons incubated with media of CRH-treated glial cells, which were prevented by administration of CXCL5/LIX receptor(CXCR2) antagonists SB225610. 7. CRHR1 activated of intracellular Gi protein to inhibit downstream AC/c AMP signaling pathway and activate Gq/PLC/PKC signaling pathway to inhibit the secretion of CXCL5/LIX in glias.(II) Regulation of synaptic formation by corticotropin-releasing hormone receptor 2 in hippocampus.(1) UCN2 but not UCN3 enhances presynaptic and postsynaptic protein expression in hippocampal slices, which was reversed by CRHR2 antagonist astressin 2B.(2) In isolated hippocampal neurons, UCN2 decreased the numbers of synapsin I- and PSD95-labeled terminals/clusters via CRHR2.(3) UCN2 enhanced the numbers and level of synapsin I- and PSD95-labeled terminals in neuron-astrocyte co-cultures.(4) Treatment of hippocampal neurons with the media of UCN2-treated astrocytes led to an increase in synapsin I- and PSD95-labeled terminals.(5) UCN2 but not UCN3 treatment induced nerve growth factor(NGF) production in astrocytes via CRHR2.(6) The effects of the media of UCN2-treated glial cells on synaptic formation in hippocampal neurons were prevented by administration of NGF receptor antagonists Trk AIg G or K252 a.Conclusions: 1. Prenatal excess GC exposure impaired spatial learning and memory and synaptic formation in adult offspring which reversed by neonatal repeated blockade of CRHR1 by CRHR1 antagonist CP154526 or antalarmin. In vitro, CRH suppresses synapsin I- and PSD95-labeled synaptic terminals formation in hippocampus via inhibition of CXCL5/LIX secretion from glial cells. 2. UCN2 but not UCN3 promotes the synaptic formation in hipppocampal neurons via induction of NGF production by astrocytes... |
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