Anti-hCG monoclonal antibody, clone D73H75

Although the inositol 1,4,5-trisphosphate (IP3) induced nuclear Ca2+ releases have been shown to play key roles in nuclear functions, the presence and operation of the IP3-dependent Ca2+ control mechanism in the nucleoplasm have not been shown. Recently, we found the presence of a high-capacity, low-affinity Ca2+-storage protein chromogranin B (CGB) and all three IP3 receptor (IP3R) isoforms in the nucleoplasm, localizing widely in both the heterochromatin and euchromatin regions. In view of the essential role of CGB-IP3R coupling in IP3-dependent Ca2+ release in the endoplasmic reticulum, the potential coupling between CGB and the IP(3)Rs in the nucleoplasm was investigated. Hence, we found in the present study the presence of a nucleoplasmic complex, which is composed of the IP3R, CGB, and phospholipids, with an estimated molecular mass of similar to 2-3 x 10(7) Da, suggesting the possibility of the presence of an IP3-sensitive Ca2+ store in the nucleoplasm. Moreover, double-labeling immunogold electron microscope studies showed the colocalization of all three IP3R isoforms with CGB to the extent that the majority of each IP3R isoform-labeling gold particles found in the nucleoplasm was literally next to the CGB-labeling gold particles. In line with the potential existence of an IP3-dependent vesicular nucleoplasmic Ca2+ store, our preliminary results indeed showed a sudden release of Ca2+ from a putative nucleoplasmic Ca2+ store in response specifically to IP3 but not to inositol 1,4-bisphosphate or inositol 1,3,4,5-tetrakisphosphate.

Experimental damage of cerebral cortex induces a slow-moving depolarization and subsequent depression of activity called cortical spreading depression (CSD) which is associated with various ionic, metabolic and genomic changes. Chromogranins are a family of water-soluble acidic proteins with a widespread distribution in secretory, large dense-core vesicles of neurons. We have earlier reported that secretogranin II (SgII) mRNA is increased in cerebral cortex hours after a unilateral craniotomy which would have induced CSD. To investigate further the regulation of chromogranin systems and the nature of genomic and biochemical changes produced by CSD, this study examined the temporal changes in chromogranin A (CgA), chromogranin B (CgB) and SgII mRNAs and CgB and SgII immunoreactivity (IR) in cerebral cortex and hippocampus following unilateral KCI-induced CSD. For comparison, the levels of mRNA for synapsin I, a protein present in small synaptic vesicles was also examined. Rats were killed at various times after 10 min or 2 h of CSD and levels of chromogranins mRNAs were determined by semiquantitative in situ hybridization histochemistry, while changes in corresponding peptide products were detected by immunohistochemistry. CSD increased both SgII and CgB mRNA levels in ipsilateral cortex - levels of SgII mRNA were significantly (P < 0.01) increased at 1-6 h after CSD (165-225% of levels in contralateral cortex), but were not significantly above control values at later time points. Increased expression of CgB mRNA was delayed and prolonged compared with SgII and was significantly (P < 0.05) increased between 3 and 24 h (120-145%) after CSD, peaked at 2 days (180%), and was still elevated at 1 week (130%) compared with contralateral cortex. No alteration in CgA mRNA was observed in the ipsilateral cortex of the same animals across the entire time-course except for an increase in piriform cortex at 1-2 days. In contrast, levels of synapsin I mRNA in affected cortex were identical to those in contralateral cortex and cortex in sham-operated rats, at all times after CSD. Levels of chromogranin (SN-IR and PE-11-IR) were also increased in ipsilateral cortex following CSD. A strong increase in SN-IR in neuronal cell bodies and fibres was observed at 12 h and a moderate increase in PE-11-IR was observed 24-72 h after CSD. These results demonstrate that chromogranin transcripts and gene products are differentially regulated by neuronal depolarization/depression occurring during CSD and suggest that these chromogranin proteins may have differing functional roles in peptide transmitter release and distinct effects on neuronal function in rat brain.