Hypothalamic astroglial connexins are required for brain glucose sensing-induced insulin secretion.

Hypothalamic glucose detection participates in maintaining glycemic balance, food intake, and thermogenesis. Although hypothalamic neurons are the executive cells involved in these responses, there is increasing evidence that astrocytes participate in glucose sensing (GS); however, it is unknown whether astroglial networking is required for glucose sensitivity. Astroglial connexins 30 and 43 (Cx30 and Cx43) form hexameric channels, which are apposed in gap junctions, allowing for the intercellular transfer of small molecules such as glucose throughout the astroglial networks. Here, we hypothesized that hypothalamic glucose sensitivity requires these connexins. First, we showed that both Cxs are enriched in the rat hypothalamus, with highly concentrated Cx43 expression around blood vessels of the mediobasal hypothalamus (MBH). Both fasting and high glycemic levels rapidly altered the protein levels of MBH astroglial connexins, suggesting cross talk within the MBH between glycemic status and the connexins' ability to dispatch glucose. Finally, the inhibition of MBH Cx43 (by transient RNA interference) attenuated hypothalamic glucose sensitivity in rats, which was demonstrated by a pronounced decreased insulin secretion in response to a brain glucose challenge. These results illustrate that astroglial connexins contribute to hypothalamic GS.

Expression profiling of prospero in the Drosophila larval chemosensory organ: Between growth and outgrowth.

BACKGROUND: The antenno-maxilary complex (AMC) forms the chemosensory system of the Drosophila larva and is involved in gustatory and olfactory perception. We have previously shown that a mutant allele of the homeodomain transcription factor Prospero (prosVoila1, V1), presents several developmental defects including abnormal growth and altered taste responses. In addition, many neural tracts connecting the AMC to the central nervous system (CNS) were affected. Our earlier reports on larval AMC did not argue in favour of a role of pros in cell fate decision, but strongly suggested that pros could be involved in the control of other aspect of neuronal development. In order to identify these functions, we used microarray analysis of larval AMC and CNS tissue isolated from the wild type, and three other previously characterised prospero alleles, including the V1 mutant, considered as a null allele for the AMC. RESULTS: A total of 17 samples were first analysed with hierarchical clustering. To determine those genes affected by loss of pros function, we calculated a discriminating score reflecting the differential expression between V1 mutant and other pros alleles. We identified a total of 64 genes in the AMC. Additional manual annotation using all the computed information on the attributed role of these genes in the Drosophila larvae nervous system, enabled us to identify one functional category of potential Prospero target genes known to be involved in neurite outgrowth, synaptic transmission and more specifically in neuronal connectivity remodelling. The second category of genes found to be differentially expressed between the null mutant AMC and the other alleles concerned the development of the sensory organs and more particularly the larval olfactory system. Surprisingly, a third category emerged from our analyses and suggests an association of pros with the genes that regulate autophagy, growth and insulin pathways. Interestingly, EGFR and Notch pathways were represented in all of these three functional categories. We now propose that Pros could perform all of these different functions through the modulation of these two antagonistic and synergic pathways. CONCLUSIONS: The current data contribute to the clarification of the prospero function in the larval AMC and show that pros regulates different function in larvae as compared to those controlled by this gene in embryos. In the future, the possible mechanism by which Pros could achieve its function in the AMC will be explored in detail.

Spatio-temporal expression of Prospero is finely tuned to allow the correct development and function of the nervous system in Drosophila melanogaster.

Adaptive animal behaviors depend upon the precise development of the nervous system that underlies them. In Drosophila melanogaster, the pan-neural prospero gene (pros), is involved in various aspects of neurogenesis including cell cycle control, axonal outgrowth, neuronal and glial cell differentiation. As these results have been generally obtained with null pros mutants inducing embryonic lethality, the role of pros during later development remains poorly known. Using several pros-Voila (prosV) alleles, that induce multiple developmental and behavioral anomalies in the larva and in adult, we explored the relationship between these phenotypes and the variation of pros expression in 5 different neural regions during pre-imaginal development. We found that the quantity of pros mRNA spliced variants and of Pros protein varied between these alleles in a tissue-specific and developmental way. Moreover, in prosV1 and prosV13 alleles, the respective decrease or increase of pros expression, affected (i) neuronal and glial cell composition, (ii) cell proliferation and death and (iii) axonal-dendritic outgrowth in a stage and cellular context dependant way. The various phenotypic consequences induced during development, related to more or less subtle differences in gene expression, indicate that Pros level needs a precise and specific adjustment in each neural organ to allow its proper function.

Targeted calretinin expression in granule cells of calretinin-null mice restores normal cerebellar functions.

Ca2 binding proteins such as calretinin, characterized by the presence of EF-hand motifs that bind Ca2+ ions, are involved in the shaping of intraneuronal Ca2+ fluxes. In the cerebellar cortex, information processing tightly relies on variations in intracellular Ca2+ concentration in Purkinje and granule cells. Calretinin-deficient (Cr-/-) mice present motor discoordination, suggesting cellular and network cerebellar dysfunctions. To determine the cell specificity of these alterations, we constructed transgenic Cr-/- mice exhibiting a selective reexpression of calretinin in granule cells through the promoter function of the GABAA receptor alpha6 subunit gene. Normal granule cell excitability and wild-type Purkinje cell firing behavior in awake mice were restored while the emergence of high-frequency oscillations was abolished. Behavioral analysis of these calretinin-rescue mice revealed that normal motor coordination was restored as compared with Cr-/- mice. These results demonstrate that calretinin is required specifically in granule cells for correct computation in the cerebellar cortex and indicate that the finetuning of granule cell excitability through regulation of Ca2+ homeostasis plays a crucial role for information coding and storage in the cerebellum.

Expression of Ca(2+) Transport Genes in Platelets and Endothelial Cells in Hypertension.

-Altered Ca(2+) handling is observed in different cells in essential hypertension. We investigated the expression of sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA) and inositol 1,4,5-trisphosphate receptor (IP(3)R) isoforms in platelets and aortic endothelial cells (EC) isolated from spontaneously hypertensive (SHR) and Wistar-Kyoto (WKY) rats by ratio reverse-transcriptase-polymerase chain reaction (RT-PCR) analysis and Western blotting. SERCA2b and SERCA3 were assessed at mRNA (EC and platelets) and at protein level (platelets). IP(3)R1, IP(3)R2, and IP(3)R3 mRNAs were demonstrated in both cell types, but only IP(3)R1 and IP(3)R2 proteins were detected in platelets. Compared with WKY, SHR EC and platelets showed higher SERCA3 and IP(3)R2 expression and lower IP(3)R1 expression. We then investigated the effect of lisinopril (20 mg. kg(-)(1). d(-)(1); 10-week treatment of 4-week-old rats or 2-week treatment of adult rats) and captopril (100 mg. kg(-)(1). d(-)(1); 2-week treatment of adult rats). Consequently, expression patterns of SERCAs and IP(3)Rs were significantly modified. Except for SERCAs mRNA in platelets, all differences between SHR and WKY disappeared. However, SERCA3 remained the predominant isoform. Both EC and platelets demonstrated a high equal expression of IP(3)R2 mRNA. IP(3)R1 was the predominant platelet protein isoform, as it was in untreated WKY. mRNA was also isolated from pancreatic islets of WKY and SHR, but no effect of either rat strain or of lisinopril treatment was observed on the expression of the studied genes. We hypothesize that the identical expression pattern of SERCAs and IP(3)Rs after treatment with ACE inhibitors represents a different nonhypertensive configuration, which, through changes in intracellular Ca(2+) handling, improves endothelial and platelet dysfunction in SHR but has no effect in WKY.