Activation of PAC1 Receptors in Rat Cerebellar Granule Cells Stimulates Both Calcium Mobilization from Intracellular Stores and Calcium Influx through N-Type Calcium Channels.

International audienceHigh concentrations of pituitary adenylate cyclase-activating polypeptide (PACAP) and a high density of PACAP binding sites have been detected in the developing rat cerebellum. In particular, PACAP receptors are actively expressed in immature granule cells, where they activate both adenylyl cyclase and phospholipase C. The aim of the present study was to investigate the ability of PACAP to induce calcium mobilization in cerebellar granule neurons. Administration of PACAP-induced a transient, rapid, and monophasic rise of the cytosolic calcium concentration ([Ca(2+)]i), while vasoactive intestinal peptide was devoid of effect, indicating the involvement of the PAC1 receptor in the Ca(2+) response. Preincubation of granule cells with the Ca(2+) ATPase inhibitor, thapsigargin, or the d-myo-inositol 1,4,5-trisphosphate (IP3) receptor antagonist, 2-aminoethoxydiphenyl borate, markedly reduced the stimulatory effect of PACAP on [Ca(2+)]i. Furthermore, addition of the calcium chelator, EGTA, or exposure of cells to the non-selective Ca(2+) channel blocker, NiCl2, significantly attenuated the PACAP-evoked [Ca(2+)]i increase. Preincubation of granule neurons with the N-type Ca(2+) channel blocker, ω-conotoxin GVIA, decreased the PACAP-induced [Ca(2+)]i response, whereas the L-type Ca(2+) channel blocker, nifedipine, and the P- and Q-type Ca(2+) channel blocker, ω-conotoxin MVIIC, had no effect. Altogether, these findings indicate that PACAP, acting through PAC1 receptors, provokes an increase in [Ca(2+)]i in granule neurons, which is mediated by both mobilization of calcium from IP3-sensitive intracellular stores and activation of N-type Ca(2+) channel. Some of the activities of PACAP on proliferation, survival, migration, and differentiation of cerebellar granule cells could thus be mediated, at least in part, through these intracellular and/or extracellular calcium fluxes

Intermittent hypoxia in a mouse model of apnea of prematurity leads to a retardation of cerebellar development and long-term functional deficits

International audienceBackground: Apnea of prematurity (AOP) is caused by respiratory control immaturity and affects nearly 50% of premature newborns. This pathology induces perinatal intermittent hypoxia (IH), which leads to neurodevelopmental disorders. The impact on the brain has been well investigated. However, despite its functional importance and immaturity at birth, the involvement of the cerebellum remains poorly understood. Therefore, this study aims to identify the effects of IH on cerebellar development using a mouse model of AOP consisting of repeated 2-min cycles of hypoxia and reoxygenation over 6 h and for 10 days starting on postnatal day 2 (P2).Results At P12, IH-mice cerebella present higher oxidative stress associated with delayed maturation of the cerebellar cortex and decreased dendritic arborization of Purkinje cells. Moreover, mice present with growth retardation and motor disorders. In response to hypoxia, the developing cerebellum triggers compensatory mechanisms resulting in the unaltered organization of the cortical layers from P21 onwards. Nevertheless, some abnormalities remain in adult Purkinje cells, such as the dendritic densification, the increase in afferent innervation, and axon hypomyelination. Moreover, this compensation seems insufficient to allow locomotor recovery because adult mice still show motor impairment and significant disorders in spatial learning.Conclusions All these findings indicate that the cerebellum is a target of intermittent hypoxia through alterations of developmental mechanisms leading to long-term functional deficits. Thus, the cerebellum could contribute, like others brain structures, to explaining the pathophysiology of AOP

A Peptidomic Approach to Characterize Peptides Involved in Cerebellar Cortex Development Leads to the Identification of the Neurotrophic Effects of Nociceptin

International audienceThe cerebellum is a brain structure involved in motor and cognitive functions. The development of the cerebellar cortex (the external part of the cerebellum) is under the control of numerous factors. Among these factors, neuropeptides including PACAP or somatostatin modulate the survival, migration and/or differentiation of cerebellar granule cells. Interestingly, such peptides contributing to cerebellar ontogenesis usually exhibit a specific transient expression profile with a low abundance at birth, a high expression level during the developmental processes, which take place within the first two postnatal weeks in rodents, and a gradual decline toward adulthood. Thus, to identify new peptides transiently expressed in the cerebellum during development, rat cerebella were sampled from birth to adulthood, and analyzed by a semi-quantitative peptidomic approach. A total of 33 peptides were found to be expressed in the cerebellum. Among these 33 peptides, 8 had a clear differential expression pattern during development, 4 of them i.e. cerebellin 2, nociceptin, somatostatin and VGF [353-372], exhibiting a high expression level during the first two postnatal weeks followed by a significative decrease at adulthood. A focus by a genomic approach on nociceptin, confirmed that its precursor mRNA is transiently expressed during the first week of life in granule neurons within the internal granule cell layer of the cerebellum, and showed that the nociceptin receptor is also actively expressed between P8 and P16 by the same neurons. Finally, functional studies revealed a new role for nociceptin, acting as a neurotrophic peptide able to promote the survival and differentiation of developing cerebellar granule neurons