ABSTRACT
Calcineurin (CaN) immunoreactivity and content increased markedly in kindled rat brain, and this increment was due to CaN in the membrane fraction. Investigation of the effects of cyclosporin A and FK506 (immunosuppressants which inhibit CaN activity in T lymphocytes) in the kindling phenomena showed that the kindling stage progression was reversibly blocked by these drugs. These findings suggest that calcineurin may play an essential role in acquiring epileptogenesis in kindling.
Subject(s)
Amygdala/physiology , Calmodulin-Binding Proteins/antagonists & inhibitors , Epilepsy/prevention & control , Immunosuppressive Agents/pharmacology , Kindling, Neurologic/physiology , Phosphoprotein Phosphatases/antagonists & inhibitors , Animals , Calcineurin , Calmodulin-Binding Proteins/metabolism , Cerebral Cortex/drug effects , Cerebral Cortex/metabolism , Cyclosporine/pharmacology , Densitometry , Epilepsy/physiopathology , Hippocampus/drug effects , Hippocampus/metabolism , Immunohistochemistry , Male , Phosphoprotein Phosphatases/metabolism , Rats , Rats, Sprague-Dawley , Tacrolimus/pharmacologyABSTRACT
The expression of a regulatory subunit of calcineurin (CaN beta) during rat spermatogenesis was examined in rat testes using a monoclonal antibody Va1. Results showed that a testis-specific isoform of CaN beta was expressed only 3 weeks after birth, when meiosis begins, and increased in amount depending on the maturation of spermatogenesis. The matured sperm, which consists of only post-meiotic cells, is most likely to have only the testis-specific isoform of CaN beta. The brain type isoform of CaN beta was not detected in rat sperm. Immunoblot analysis of testes from different rodent species by a monoclonal antibody Va1 showed that all rodent species examined had their own homologues corresponding to a testis-specific isoform of CaN beta in rats, although they showed distinctively different molecular weights on SDS-PAGE compared to the testis-specific isoform in rats. Each homologue was shown to be specifically expressed in post-meiotic phase of spermatogenesis, as was seen in rats.
Subject(s)
Aging/metabolism , Antibodies, Monoclonal , Calmodulin-Binding Proteins/metabolism , Isoenzymes/metabolism , Meiosis , Phosphoprotein Phosphatases/metabolism , Spermatogenesis/physiology , Testis/metabolism , Animals , Brain/growth & development , Brain/metabolism , Calcineurin , Calmodulin-Binding Proteins/chemistry , Electrophoresis, Polyacrylamide Gel , Guinea Pigs , Immunoblotting , Isoenzymes/chemistry , Macromolecular Substances , Male , Mice , Molecular Weight , Phosphoprotein Phosphatases/chemistry , Rats , Testis/growth & developmentABSTRACT
Identification of a src-related tyrosine kinase and its regional and cellular distributions were studied in rat brain. The study was performed using a specific antibody raised against a synthetic peptide corresponding to the conserved autophosphorylation site of src-family tyrosine kinases. The antibody (alpha-src antibody) recognized a 43 kDa polypeptide in plasma membrane enriched fraction. The immunohistochemical data revealed that the polypeptide is localized in astrocytes of corpus callosum and fimbria hippocampus. The presence of this src-related protein in astrocytes suggests that it may have some control in their proliferation and differentiation.
Subject(s)
Astrocytes/metabolism , Proto-Oncogene Proteins pp60(c-src)/metabolism , Animals , Brain/cytology , Brain/metabolism , Fluorescent Antibody Technique , Glial Fibrillary Acidic Protein/metabolism , Immunohistochemistry , Rats , Rats, Inbred Strains , Tissue DistributionABSTRACT
Two isoforms of calcineurin beta subunit(beta 1 and beta 2) were identified in rat testis by a monoclonal antibody Va1. Both beta 1 and beta 2 were recovered in calmodulin binding protein fraction and showed calcium shift on SDS-polyacrylamide gel electrophoresis which is the specific character for EF-hand calcium binding protein. beta 2 showed same apparent molecular weight on SDS-PAGE as that of brain calcineurin beta and was found in wide variety of tissues. beta 1 was shown to have six amino acid polypepeptide sequence and it showed higher molecular weight than brain beta and was specific for testis.
Subject(s)
Calmodulin-Binding Proteins/analysis , Isoenzymes/analysis , Phosphoprotein Phosphatases/analysis , Testis/enzymology , Amino Acid Sequence , Animals , Antibodies, Monoclonal , Calcineurin , Calmodulin-Binding Proteins/isolation & purification , Electrophoresis, Polyacrylamide Gel , Immunoblotting , Macromolecular Substances , Male , Molecular Sequence Data , Oligopeptides/chemical synthesis , Oligopeptides/isolation & purification , RatsABSTRACT
We have cloned and sequenced rat testis cDNAs coding for a calcium binding polypeptide similar to calcineurin beta subunit, the Ca(2+)-binding subunit of the Ca2+/calmodulin stimulated protein phosphatase. Rat testis cDNA library was screened with a monoclonal antibody Va1 raised against bovine brain calcineurin beta subunit. The deduced amino acid sequence is similar to that of human brain calcineurin beta subunit with respect to containing four putative calcium binding sites. However, distinct differences were found: 1) The cloned cDNA had six amino acids polypeptide tail at carboxy-terminal which is absent in human brain calcineurin beta subunit. This amino acids tail makes the carboxy-terminal highly hydrophilic in contrast to the human brain beta subunit which is hydrophobic at carboxy-terminal; 2) eleven amino acids at the N terminal of the cloned cDNA were completely different from the corresponding region of the brain calcineurin beta subunit.
