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Biomedical subjects

A J Silva

Publications and source records attributed to A J Silva.

At least 19 recordsLinked to original sources
PubMed1996

Ischemia-induced neuronal damage: a role for calcium/calmodulin-dependent protein kinase II.

Calcium/calmodulin-dependent protein kinase II (CaM-kinase) is a central enzyme in regulating neuronal processes. Imbalances in the activity and distribution of this enzyme have been reported following in vivo ischemia, and sustained decreases in activity correlate with subsequent neuronal death. In this report, mice that had been rendered deficient in the alpha subunit of CaM-kinase using gene knock-out technology were utilized to determine whether this enzyme is causally related to ischemic damage. Using a focal model of cerebral ischemia, we showed that homozygous knock-out mice lacking the alpha subunit exhibited an infarct volume almost twice that of wild-type litter mates. Heterozygous mice exhibited slightly less damage following ischemia than did homozygous mice, but infarct volumes remained significantly larger than those of wild-type litter mates. We conclude that reduced amounts of the alpha subunit of CaM-kinase predisposes neurons to increased damage following ischemia and that any perturbation that decreases the amount or activity of the enzyme will produce enhanced susceptibility to neuronal damage.

Actins↗
PubMed1995

Limbic epilepsy in transgenic mice carrying a Ca2+/calmodulin-dependent kinase II alpha-subunit mutation.

Multifunctional Ca2+/calmodulin-dependent protein kinase II (CaMK) phosphorylates proteins pivotally involved in diverse neuronal processes and thereby coordinates cellular responses to external stimuli that regulate intracellular Ca2+ [Hanson, P. I. & Schulman, H. (1992) Annu. Rev. Biochem. 61, 559-664]. Despite extensive study, the impact of this enzyme on control of the excitability of neuron populations in the mammalian nervous system in situ is unknown. To address this question, we studied transgenic mice carrying a null mutation (-/-) for the alpha subunit of CaMK. In contrast to wild-type littermates, null mutants exhibit profound hyperexcitability, evident in epileptic seizures involving limbic structures including the hippocampus. No evidence of increased excitability was detected in mice carrying null mutations of the gamma isoform of protein kinase C, underscoring the specificity of the effect of CaMK. CaMK plays a powerful and previously underappreciated role in control of neuronal excitability in the mammalian nervous system. These insights have important implications for analyses of mechanisms of epilepsy and, perhaps, learning and memory.

Animals↗
PubMed1995

The alpha-Ca2+/calmodulin kinase II: a bidirectional modulator of presynaptic plasticity.

The alpha-Ca2+/calmodulin kinase II (alpha CaMKII) is required for long-term potentiation in the CA1 region of the hippocampus. Here, we report that this kinase also has a crucial role in presynaptic plasticity. Paired-pulse facilitation is blunted in the CA1 region of mice heterozygous for a targeted mutation of alpha CaMKII, confirming that this kinase can promote neurotransmitter release. Unexpectedly, field and whole-cell recordings of posttetanic potentiation show that the synaptic responses of mutants are larger than those of controls, indicating that alpha CaMKII can also inhibit transmitter release immediately after tetanic stimulation. Thus, alpha CaMKII has the capacity either to potentiate or to depress excitatory synaptic transmission depending on the pattern of presynaptic activation.

Animals↗
PubMed1994

Deficient long-term memory in mice with a targeted mutation of the cAMP-responsive element-binding protein.

The cAMP-responsive element-binding protein (CREB) has been implicated in the activation of protein synthesis required for long-term facilitation, a cellular model of memory in Aplysia. Our studies with fear conditioning and with the water maze show that mice with a targeted disruption of the alpha and delta isoforms of CREB are profoundly deficient in long-term memory. In contrast, short-term memory, lasting between 30 and 60 min, is normal. Consistent with models claiming a role for long-term potentiation (LTP) in memory, LTP in hippocampal slices from CREB mutants decayed to baseline 90 min after tetanic stimulation. However, paired-pulse facilitation and posttetanic potentiation are normal. These results implicate CREB-dependent transcription in mammalian long-term memory.

Animals↗
PubMed1994

Targeting learning.

Novel transgenic approaches provide an exciting opportunity to assess the impact of the loss of specific genes in the biochemistry and electrophysiology of neurons involved in a learned behavior. Recent studies describing mice harboring mutations in five kinase genes expressed in the hippocampus found that two of these kinases, the alpha-Ca(2+)-calmodulin-dependent kinase II and the Fyn tyrosine kinase are necessary for the establishment of long-term potentiation. In addition to providing a new tool for the dissection of the molecular mechanisms of synaptic plasticity, these mutants will be important in determining how changes in synaptic strength affect not only learning and memory, but also a host of other processes thought to be associated with plasticity.

Animals↗
PubMed1994

Plastic genes are in!

Even though the synthesis of new proteins is thought to be essential for long-term changes in synaptic plasticity, as well as for long-term memory, little is known about the identity of the required proteins. The hunt for these molecules is under way, however, and in the past year several groups of researchers have entered this fascinating search by introducing new approaches that have lead to the identification of several potential candidates, amongst which are trophic factors, kinases, ion channels, and proteases. The results will have much to say not only about the nature of memory, but also about the mechanisms of learning.

Animals↗
PubMed1993

Modified hippocampal long-term potentiation in PKC gamma-mutant mice.

Calcium-phospholipid-dependent protein kinase (PKC) has long been suggested to play an important role in modulating synaptic efficacy. We have created a strain of mice that lacks the gamma subtype of PKC to evaluate the significance of this brain-specific PKC isozyme in synaptic plasticity. Mutant mice are viable, develop normally, and have synaptic transmission that is indistinguishable from wild-type mice. Long-term potentiation (LTP), however, is greatly diminished in mutant animals, while two other forms of synaptic plasticity, long-term depression and paired-pulse facilitation, are normal. Surprisingly, when tetanus to evoke LTP was preceded by a low frequency stimulation, mutant animals displayed apparently normal LTP. We propose that PKC gamma is not part of the molecular machinery that produces LTP but is a key regulatory component.

Animals↗
PubMed1993

Mosaic methylation in clonal tissue.

Current models suggest that de novo methylases add methyl groups to mammalian DNA early in development, establishing cell-specific patterns of methylation, and that these patterns are maintained by maintenance methylases that copy them onto newly replicated DNA strands. To test the prediction that clonal populations of histologically homogeneous cells should, therefore, have homogeneous methylation patterns, we studied methylation in leiomyomas. Despite the clonality and histological homogeneity of these solid tumors, we found that cells were heterogeneously methylated at a number of genomic sites. The heterogeneity was not caused by random methylation events within the leiomyomas because methylation patterns were similar in the core and periphery of a given tumor, and similar also among samples of independent leiomyomas and surrounding myometrial tissues extracted from a single uterus. Our results also showed that methylation of a site in the YNZ22 locus--in leiomyomas and in smooth muscle--was determined independently from the methylation of a neighboring site. Similar results were obtained for the IGH locus in colon and in several tumor tissues. These data indicate that methylation patterns are not identical in progeny cells, as current models suggest. Instead, it seems likely that methylation of a specific site reflects an equilibrium frequency defined by a continual loss and gain of methyl groups. Hence, the specificity found for the methylation of mammalian tissues is not achieved by strictly determining the methylation fate of individual cells, but by determining the overall methylation frequencies for individual sites.

Cell Differentiation↗
PubMed1992

Impaired spatial learning in alpha-calcium-calmodulin kinase II mutant mice.

Although long-term potentiation (LTP) has been studied as the mechanism for hippocampus-dependent learning and memory, evidence for this hypothesis is still incomplete. The mice with a mutation in the alpha-calcium-calmodulin-dependent kinase II (alpha-CaMKII), a synaptic protein enriched in the hippocampus, are appropriate for addressing this issue because the hippocampus of these mice is deficient in LTP but maintains intact postsynaptic mechanisms. These mutant mice exhibit specific learning impairments, an indication that alpha-CaMKII has a prominent role in spatial learning, but that it is not essential for some types of non-spatial learning. The data considerably strengthen the contention that the synaptic changes exhibited in LTP are the basis for spatial memory.

Animals↗
PubMed1992

Deficient hippocampal long-term potentiation in alpha-calcium-calmodulin kinase II mutant mice.

As a first step in a program to use genetically altered mice in the study of memory mechanisms, mutant mice were produced that do not express the alpha-calcium-calmodulin-dependent kinase II (alpha-CaMKII). The alpha-CaMKII is highly enriched in postsynaptic densities of hippocampus and neocortex and may be involved in the regulation of long-term potentiation (LTP). Such mutant mice exhibited mostly normal behaviors and presented no obvious neuroanatomical defects. Whole cell recordings reveal that postsynaptic mechanisms, including N-methyl-D-aspartate (NMDA) receptor function, are intact. Despite normal postsynaptic mechanisms, these mice are deficient in their ability to produce LTP and are therefore a suitable model for studying the relation between LTP and learning processes.

Animals↗
PubMed1988

Inheritance of allelic blueprints for methylation patterns.

We have developed a strategy to distinguish between the methylation patterns of homologous chromosomes in tissues, and to follow these patterns in human pedigrees. This genetic approach uncovered evidence of variation in the methylation of allelic sites on homologous chromosomes. This variation was tissue-specific and reproducible after transmission through the germ line, demonstrating that homologous chromosomes have distinct blueprints for the tissue-specific regulation of methylation. Furthermore, this approach can be used to study the relationship between parental imprinting and methylation in native mammalian loci.

Alleles↗
PubMed1987

Characterization of a highly polymorphic region 5' to JH in the human immunoglobulin heavy chain.

A cloned DNA segment 1.25 kilobases (kb) upstream from the joining segments of the human heavy chain immunoglobulin gene revealed extensive polymorphic variation at this locus, and the polymorphic pattern was stably transmitted to the next generation. Genomic restriction analysis showed that the polymorphism was caused by insertions/deletions within an MspI/BamHI fragment. Sequencing of one allele, 848 base pairs (bp) long, revealed eleven 50-base-pair tandem repeats. A second allele, 648 bp long, was cloned from a human genomic cosmid library, sequenced, and found to contain four fewer repeats than the first allele. A survey of 186 chromosomes from unrelated individuals of primarily northern European descent revealed at least six alleles.

Alleles↗