Expression and subcellular localization of multifunctional calmodulin-dependent protein kinases-I, -II and -IV are altered in rat hippocampal CA1 neurons after induction of long-term potentiation.

Long-term potentiation (LTP) is considered to be associated with an increase in expression as well as activity of Ca(2+)/calmodulin-dependent protein kinases (CaMKs). LTP-induced and control hippocampal slices were studied by immunohistochemical and electronmicroscopic analyses using anti-CaMK-I, -II and -IV antibodies. All three kinases were demonstrated to increase their expression in CA1 neurons. CaMK-I was shown to mainly localize in the cytoplasm of the control and LTP-induced neurons, and a significant increase of immunoreactivity was observed in the latter neurons. A part of CaMK-I was found to translocate to the nuclei of LTP-induced hippocampal CA1 neurons. Direct evidence of the translocation of CaMK-II from cytoplasm to nuclei in LTP was demonstrated by immuno-electronmicroscopy. A significant increase in expression of CaMK-IV in the nuclei was also observed. Our data suggest that all the three CaMKs were actively involved in nuclear Ca(2+)-signaling in LTP.

Reticalmin: a novel calcium/calmodulin-dependent protein kinase IV-like protein in rat retina.

Western blot analysis of 100,000 g supernatant of rat retina using a polyclonal anti-Ca2+/ calmodulin-dependent protein kinase IV (CaM-kinase IV) antibody revealed an immunoreactive mass of 35 kDa, termed reticalmin. Lower amount of a isoform of CaM-kinase IV was also expressed in rat retina. Reticalmin did not react with anti-CaM-kinase IV C-terminal peptide antibody which recognized alpha and beta isoforms of CaM-kinase IV and calspermin. Immunohistochemically reticalmin was shown to be localized mainly in the outer segment of photo-receptor cells, and in dendrites of inner plexiform layers and may be in nuclei of ganglion cells and some inner nuclear layer cells.

Involvement of calmodulin-dependent protein kinases-I and -IV in long-term potentiation.

Multifunctional Ca2+/calmodulin-dependent protein kinases (CaMKs) are thought to be involved in the induction of long-term potentiation (LTP). In the present study, LTP was induced by theta burst stimulation in the Schaffer collateral area of the stratum radiatum in the hippocampal CA1 region of the rat hippocampus. LTP-induced and control hippocampal slices were studied by Western blot and immunohistochemical analyses using CaMK-I, -II and -IV antibodies. Increased amounts of all three CaMKs were found in LTP-induced hippocampal slices as indicated by Western blot as well as by the density of their immunoreactivity. Our data clearly shows that not only CaMK-II but also CaMK-I and -IV contribute to synaptic plasticity formed in LTP.

A chronological study of the expression of glial fibrillary acidic protein and calbindin-D28 k by reactive astrocytes in the electrically lesioned rat brain.

Immunoreactivity of neuronal and glial marker proteins of reactive astrocytes around the electrically damaged pyramidal layer and stratum radiatum of the hippocampal CA1 region and corpus callosum was chronologically studied in electrically lesioned rat brains. A monoclonal antibody against calbindin-D28 k (CD28-Ab) and a polyclonal antibody against glial fibrillary acidic protein (GFAP-Ab) were used for immunostaining. Immunoreactivity of CD28 and GFAP in the reactive astrocytes was detected in brains 1-6 weeks post-lesion but not in non-lesioned brains. The number of immunohistochemically stained reactive astrocytes around the electrically damaged areas were counted and then compared with the number of those in the same areas of non-lesioned brains. The number of CD28- and GFAP-immunoreactive astrocytes began to increase around the lesion from 1-3 weeks following lesion in the pyramidal layer of the hippocampal CA1 region and from 1-4 weeks following lesion in the stratum radiatum of the hippocampal CA1 region and corpus callosum. These immunoreactive astrocytes could be observed for 6 weeks (the maximum survival time studies) in all areas of the lesioned brains studied. The increase in the number of reactive astrocytes might have been induced by the stimulatory effects of neurotrophic factors, or growth factors, produced around the lesioned site. The constancy in the number of reactive astrocytes after 3 and 4 weeks in the lesioned areas may have been due to the termination of the initial phase of the repair process, i.e. space-filling. Reactive astrocytes which were stained by GFAP-Ab were separated into two groups, based on the presence of CD28, i.e. CD28-positive and CD28-negative reactive astrocytes. The presence of CD28 might confer certain functions via calcium-mediated mechanisms on CD28-positive astrocytes in addition to the constructive role mediated by GFAP.

Expression of calbindin-D28K by reactive astrocytes in gerbil hippocampus after ischaemia.

Calbindin-D28K (Calbindin) is a member of the superfamily of calcium-binding proteins that is implicated in the regulation of intracellular calcium. In the adult mammalian brain, calbindin was thought to be present only in neurones, where it is believed to serve a neuroprotective role. We now report the expression of calbindin after ischaemia in reactive astrocytes in the CA1 subfield of the hippocampus. Since other calcium-binding proteins, such as S-100 and calmodulin, which induce transformation or proliferation of glia, occur in astrocytes, it is conceivable that the expression of calbindin after ischaemia might be an important part of the process of gliosis.