Biomedical subjects

C G Wasterlain

Publications and source records attributed to C G Wasterlain.

At least 91 records · Page 5Linked to original sources

PubMed1988

Localization of retinal calmodulin kinase.

The localization of calmodulin kinase II (CaM kinase) was studied in the retina by light and electron microscopic immunocytochemistry, and by enzymatic and immunoblot assay of cellular and subcellular tissue fractions. By light microscopy, both mono- and polyclonal antibodies revealed CaM kinase-like immunoreactivity in the inner and outer plexiform regions (synaptic layers), retinal pigment epithelium (RPE), and ganglion cells. The inner nuclear layer and photoreceptor outer segments stained much less intensely, and the outer nuclear layer did not stain. Electron microscopy confirmed the high concentration of immunoreactive protein in RPE and minimal outer segment staining. In addition, photoreceptor inner segments also contained CaM kinase-like immunoreactivity. Calcium and calmodulin stimulated phosphate incorporation into proteins of retinal cytosol and of isolated and cultured RPE. Calcium- and calmodulin-dependent kinase activity was present to a lesser degree in crude nuclei and synaptic membranes and was absent in isolated rod outer segments. Immunoblot analyses were consistent with enzymatic assays and immunocytochemistry. These data suggest that retinal CaM kinase is ideally located to play an important role in synaptic transmission and modulation of visual processes. Furthermore, its presence in RPE implies that CaM kinase may have a more ubiquitous role in regulating cellular processes than was previously recognized.

Animals↗

PubMed1988

A retinal calmodulin-dependent kinase: calcium/calmodulin-stimulated and -inhibited states.

A calcium/calmodulin-dependent protein kinase was isolated from retina. The retinal enzyme is composed exclusively of 50-kilodalton (kD) subunits and has a molecular mass of approximately 275 kD, in contrast to forebrain calmodulin kinase II, which is composed of 50-kD and 60-kD subunits in a 3:1 ratio and has a molecular mass of approximately 520 kD. Similar substrate specificities, kinetic properties, capacity to bind calmodulin, and immunoreactivity suggest that the retinal kinase is an isoenzyme of forebrain calmodulin kinase II. Both kinases autophosphorylate in an intramolecular manner; however, autophosphorylation has different effects on the activities of the two enzymes. Autophosphorylation of retinal calmodulin kinase converts the enzyme from a calcium/calmodulin-dependent to a calcium/calmodulin-inhibited kinase, with high activity in the absence of calcium, whereas autophosphorylation of the forebrain kinase results in a less active, calcium/calmodulin-independent enzyme. These properties of calmodulin kinase may play an important role in retinal function.

Animals↗

PubMed1988

Low [3H]cytochalasin B binding in the cerebral cortex of newborn rat.

The concentrations of glucose transporter in the cerebral cortex and brainstem of neonatal (4-7 days old) and adult rats were measured using [3H]cytochalasin B binding. There was significantly lower binding in neonatal cortex (1.9 +/- 0.7 pmol/mg protein) compared to adult (8.9 +/- 2.5 pmol/mg protein). Scatchard analysis indicates this difference is due to a lower Bmax (neonate, 9.7 pmol/mg protein; adult, 18.6 +/- 1.3 pmol/mg protein). Measurement of [3H]cytochalasin B binding in microvessels prepared from cortex of adult (28.1 +/- 3.5 pmol/mg protein) and neonate (12.8 +/- 1.9 pmol/mg protein) indicates a lower binding in the microvasculature of neonates, whereas no such difference was seen in the binding in microvessels prepared from adult and neonatal brainstem (adult, 11.8 +/- 2.3 pmol/mg protein; neonate, 9.4 +/- 2.7 pmol/mg protein). In both adult and neonate brain, there is an enrichment of glucose transporters in the microvasculature.

Aging↗

PubMed1986

Habituation of the local cyclic GMP response during amygdaloid carbachol kindling in the rat.

Seizures kindled with amygdaloid carbachol injections are transynaptic, dependent on activation of a specific population of muscarinic receptors, and some components of their expression could be mediated by intracellular second messengers. We measured cyclic GMP and cyclic AMP concentrations in micropunch biopsies of multiple brain regions after microwave fixation during the development and the expression of carbachol-kindled seizures in the rat. In the naive carbachol-injected amygdala, cyclic GMP concentrations rose from 1.03 +/- 0.15 pmol/mg protein to 2.21 +/- 0.46 after 2 min, and significant rises occurred in caudate, hypothalamus and contralateral amygdala. This response did not occur in implanted controls, after injection of mock cerebrospinal fluid, or when carbachol actions were blocked with atropine. The rise in cyclic GMP progressively disappeared upon repeated stimulation (injected amygdala on tenth stimulation: 0.72 +/- 0.23 pmol/mg protein). However, a late rise in both cyclic GMP and cyclic AMP concentrations occurred in many brain regions during convulsive seizures. These data suggest that during the development of kindling, changes in neuronal and synaptic excitability are associated with changes in intracellular second messengers.

Action Potentials↗

PubMed1986

Preferential blood flow to brainstem during generalized seizures in the newborn marmoset monkey.

The effect of generalized seizures on local cerebral blood flow was studied autoradiographically in 21 immature marmoset monkeys, using either [123I]- or [131I]isopropyliodoamphetamine. Generalized convulsions were induced in ketamine-anesthetized and awake monkeys with bicuculline and continued for 4-59 min. During convulsions in marmosets less than 3 weeks of age, there was a striking rearrangement of blood flow in favor of the brainstem pontomedullary region. The ratios of blood flow in pons-medulla to blood flow in cerebral cortex, putamen, ventroposterior thalamic nuclei, lateral geniculate nuclei, cerebellum and hemispheric white matter increased 1 1/2 to 2 times compared to controls. In seizure animals 4-8 weeks of age, the redistribution of blood flow to brainstem did not occur. Although metabolic acidosis developed after 30 min of bicuculline-induced seizures, mean arterial blood pressure, temperature, arterial pO2 and pCO2 did not differ significantly from controls, indicating that hypoxemia, hypercapnia and hypotension cannot explain the altered cerebral blood flow pattern. The redistribution phenomenon could be explained by more pronounced vasodilatation in brainstem than many other brain regions during generalized seizures in newborn monkeys. Lack of significant vasodilatation in forebrain structures such as cerebral cortex could contribute to neuronal damage by limiting substrate supply at a time of increased metabolic activity.

Amphetamines↗

PubMed1986

Kindling induces a long-lasting change in the activity of a hippocampal membrane calmodulin-dependent protein kinase system.

Septal kindling has been shown to produce a long-lasting decrease in endogenous calcium/calmodulin-dependent phosphorylation of hippocampal synaptic plasma membrane proteins, including two major bands of approximately 50,000 and 60,000 Daltons. These two proteins differ from the B-50 protein and tubulin, as evidenced by differences in migration in SDS-PAGE gels and by lack of cross-immunoreactivity with specific antibodies. Identity of these two proteins with the rho and sigma subunits of purified calmodulin-dependent kinase (CaM Kinase II) is suggested by similar migration in SDS-PAGE and two-dimensional gels, by similar calmodulin binding in two-dimensional gels, and similar 125I-peptide mapping of the 50,000 Dalton protein. These results demonstrate that septal kindling is associated with changes in the activity of a major Ca2+/calmodulin-dependent kinase system in hippocampal synaptic plasma membrane. This long-lasting modulation of kinase activity may provide a molecular insight into some aspects of neuronal plasticity.

Animals↗

PubMed1986

Autophosphorylation of calmodulin kinase II: functional aspects.

Autophosphorylation of purified calmodulin kinase II dramatically inhibited protein kinase activity and enhanced substrate selectivity. Inhibition was observed over a wide range of calmodulin concentrations but calmodulin binding was unaffected. Autophosphorylation of calmodulin kinase II may be a mechanism for limiting phosphorylation to physiological substrates and terminating some of calcium's actions in synaptic events.

Adenosine Diphosphate↗

PubMed1986

Metabolic anatomy of generalized bicuculline seizures in the newborn marmoset monkey.

Sustained convulsive seizures were induced with bicuculline in ketamine-anesthetized marmoset monkeys aged 7 to 18 days. Relative 2-deoxyglucose metabolism was compared in convulsing (N = 9) and control (N = 6) animals. Convulsions were accompanied by striking focal increases in cerebral 2-deoxyglucose uptake which were remarkably consistent from animal to animal. Increased 2-deoxyglucose uptake in broad cortical regions (2- to 3-fold) suggests that cortical mechanisms can be important, even in neonatal seizures. The hippocampus and other limbic system structures were markedly activated, as were nuclei of the basal ganglia and thalamus. In contrast, sensory systems were less affected. No increase in 2-deoxyglucose uptake was found in the lateral geniculate nuclei, and a 22% decrease was found in the inferior colliculus (central core). Increased uptake was found in several white matter regions, and activation of the corpus callosum (2.6-fold) was comparable to that found for many gray matter regions. Our results show that generalized bicuculline seizures can produce striking focal increases in cerebral 2-deoxyglucose metabolism in brain regions known to be vulnerable to epileptic brain damage.

Animals↗

PubMed1986

Cyclic nucleotide response of the hippocampal formation to septal stimulation in naive and kindled rats.

Rats were kindled through nonmagnetic electrodes stereotaxically implanted into the medial septum. Concentrations of cyclic AMP and cyclic GMP were measured by radioimmunoassay in seven brain regions after microwave fixation during the development and expression of kindled seizures. Hippocampal concentrations were similar to untreated controls (cyclic GMP level in the left and right hippocampus, 0.66 +/- 0.04 and 0.68 +/- 0.07 pmol/mg of protein, respectively; cyclic AMP, 9.4 +/- 0.9 and 9.6 +/- 0.8 pmol/mg of protein, respectively), in kindled animals that were not stimulated, and in naive animals in response to septal stimulation, in spite of the presence in the latter group of bilateral hippocampal afterdischarges. Animals that failed to develop kindling and kindled animals that failed to have a seizure in response to stimulation also showed no change in cyclic nucleotide concentrations in any brain region. Kindled animals that developed a seizure following stimulation showed significant elevations in levels of both cyclic GMP and cyclic AMP in hippocampus and in several other brain regions. A single naive animal that had a seizure in response to its first stimulation also appeared to have elevated concentrations of both cyclic nucleotides in hippocampus. These data suggest that the elevation in levels of both cyclic GMP and cyclic AMP during kindled seizures is associated with seizure development rather than with the generation of afterdischarges or with the kindling engram.

Animals↗

PubMed1986

Effect of altered blood plasma osmolalities on regional brain amino acid concentrations and focal seizure susceptibility in the rat.

Blood plasma hypo- or hyperosmolality alters significantly the concentration of some amino acids in brain tissues of the medial septum and hippocampus of adult Sprague-Dawley rats. With some notable exceptions, brain amino acid concentrations decreased under hypoosmotic conditions and increased under hyperosmotic conditions. Osmotic changes and brain amino acid changes appear to be related to each other in an almost linear fashion. A comparison of rats and toads indicates that the patterns of changes in brain amino acid concentrations in response to a hypoosmotic plasma osmolality were almost identical for both species. Changes achievable under hyperosmotic conditions were considerably greater in toads. When rats with kindled epileptogenic foci were made hypoosmotic by water-loading, seizure thresholds decreased dramatically. Our data suggest a possible relationship between the hypoosmotically induced biochemical changes in brain tissues (especially some amino acid neurotransmitters and neurotransmitter precursors) and the hypoosmotically induced increase in seizure susceptibility.

Amino Acids↗

PubMed1986

Brain protein metabolism in epilepsy.

Both generalized and focal seizures dissociate brain polyribosomes and severely inhibit brain protein synthesis. This effect is found in freely convulsing animals and in animals that have been paralyzed and oxygen-ventilated in order to prevent hypoxemia, cerebral hypoxia, and other systemic changes associated with convulsions. Recent autoradiographic studies have shown that generalized seizures can result in striking focal inhibition of brain protein synthesis in adult rats and newborn marmoset monkeys. Local cerebral glucose metabolism and local cerebral blood flow were also studied in newborn marmosets by autoradiography. Although flow and metabolism are closely matched in control marmosets, seizures result in large local increases in 2-deoxyglucose metabolism, with lesser or no increases in local cerebral blood flow resulting in a relative mismatch. Those regions in which protein synthesis was most severely inhibited were those in which the relative mismatch between blood flow and metabolism was most marked. The molecular mechanisms regulating protein biosynthesis are not known. Translational regulation during seizures appears to be exerted, in large part, at the initiation step. A likely mechanism is the inhibition of ternary complex formation, one of the early steps in the initiation process, by increases in the intracellular ratio of [GDP]:[GTP]. This ratio is related to the cells' energy charge. Reduced levels of ATP during seizures can lead to an increased ratio of [GDP]:[GTP] via of the enzyme nucleoside diphosphate kinase (E.C. 2.7.4.6) and to inhibition of protein synthesis initiation. Regulation of protein biosynthesis during seizures is likely to be complex and exerted at many sites; some of these possibilities are discussed.

Acid-Base Equilibrium↗

PubMed1986

Synaptic mechanisms in the kindled epileptic focus: a speculative synthesis.

This chapter reviews the chemical kindling model of epilepsy and speculates on its significance. Both human and experimental epilepsies are extremely heterogeneous, and it is unlikely that a single molecular or cellular mechanism can account for such a diversity of behavioral manifestations. Recent studies of chemical kindling favor the view that in this model, epilepsy is a property of neuronal networks that can take place in a structurally intact brain and does not depend on the presence of gross or microscopic brain damage. Kindling can be obtained by daily injections of nanomolar amounts of multiple muscarinic agonists in selective brain regions such as the amygdala and, once acquired, it is very persistent and frequently accompanied by spontaneous seizures. No evidence exists for creation of a novel pathway, and studies of seizure threshold suggest the need for a critical mass of neurons even on initial stimulation. The amounts of muscarinic agents injected are small enough to have little recordable effect initially, and the number of stimulations needed varies directly with the dose and inversely with the interstimulus interval. Carbachol kindling is inhibited by picomolar amounts of muscarinic antagonists, and the relative potencies of drugs on the kindling behavior in vivo parallel their affinity for muscarinic receptors in vitro. The (+) isomer of acetyl-beta-methylcholine, with good affinity for the muscarinic receptor, can induce kindling, whereas the (-) stereo isomer with poor affinity for the receptor cannot. No morphological differences are observed between animals injected with the (+) or the (-) isomer. These experiments suggest that the development of chronic focal epilepsy can take place in a structurally intact brain, be independent of the production of brain damage, and totally dependent on synaptic excitation. In other words, in this model, epilepsy may be a disease of cell-cell communication in which structurally normal neurons develop epileptiform responses as their interactions are modified through synaptic activation. A study of the relationships between carbachol and electrical kindling of the same site gave different results depending on the site of stimulation. In the amygdala, no interaction was found, but when both stimuli were aimed at the cholinoceptive hippocampal cells, a strong facilitation in both directions was observed. Thus, it appears that chemical and electrical kindling share similar mechanisms and that cross-facilitation depends on the existence of a common anatomy. The same anticonvulsants that block electrical kindling also inhibit chemical kindling.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1985

Transfer between chemical and electrical kindling in the septal-hippocampal system.

We re-investigated the interaction between chemical and electrical kindling in two anatomical locations: the amygdaloid region and the septal-hippocampal complex. Amygdaloid animals were implanted with a chemitrode into the left basolateral amygdala, which could then be stimulated electrically (400 microA, 1 s, 60 Hz, AC) or chemically by injection of carbachol (1 microliter, 2.7 nmol, sterile, isotonic). Septal-hippocampal animals were implanted with an electrode high in the medial septum, a cannula in the dorsal hippocampus. In both groups, half the animals were kindled electrically, and after one week of rest chemical kindling was begun. The other half were kindled chemically first, then electrically. The result differed with the anatomical location. With amygdaloid implants, no significant transfer was observed. In the septal-hippocampal group, by contrast, significant interactions were observed in both directions. These results suggest that chemical and electrical kindling involves similar mechanisms, and that the extent to which transfer occurs reflects the degree to which they share a common chemical anatomy.

Amygdala↗

PubMed1985

Cholinergic kindling: what has it taught us about epilepsy?

We reviewed recent evidence that chemical kindling of epileptic seizures can be induced by injection into the amygdala of multiple cholinergic muscarinic agonists, and blocked by multiple muscarinic antagonists. The stereospecific induction of kindling by (+) but not by (-) acetyl-beta-methylcholine shows that some types of repeated synaptic activation can produce epilepsy, in the absence of specific brain damage. The failure of bicuculline (but not of carbachol) to produce kindling with amygdaloid injections, and its ability to produce a limited seizure spread in neocortex, suggest that repetitive seizure activity alone is not sufficient to produce kindling. A review of some recent neurochemical changes in the synaptic apparatus associated with some types of kindling suggests potential areas for future investigation, but no cause-and-effect relationship between neurochemical and behavioral changes can be inferred so far.

Amygdala↗

PubMed1985

Neonatal seizures in monkeys and rabbits: brain glucose depletion in the face of normoglycemia, prevention by glucose loads.

Sustained convulsive seizures were induced with bicuculline in newborn rabbits and marmoset monkeys. In both species, seizures were predominantly tonic, with generalized polyspikes on the EEG. Brain glucose concentration fell dramatically during seizures in both species, and in many normoglycemic animals reached levels usually associated with severe hypoglycemia, suggesting that glucose transport from blood to brain could not keep pace with glucose utilization. Glucose loads given to rabbits induced hyperglycemia and during seizures maintained brain glucose well above concentrations needed to saturate hexokinase so that glycolytic rates were probably never limited by substrate availability. Blood and brain lactate concentrations rose during seizures but never reached levels considered cytotoxic. These data suggest that epileptic seizures can deplete brain glucose in normoglycemic neonates of several species, including subhuman primates.

Animals↗

PubMed1984

Selective focal inhibition of brain protein synthesis during generalized bicuculline seizures in newborn marmoset monkeys.

Generalized epileptic seizures, induced in 5-7 day-old marmoset monkeys with bicuculline (5 mg/kg) produce striking focal disruption of protein synthesis. Inhibition of protein synthesis was most severe in the neocortex and hippocampus but was completely absent in the lateral geniculate nuclei and in the optic tract. In the cortex, regions in which protein synthesis was most severely reduced alternated with areas in which it was relatively preserved creating a columnar pattern. With few exceptions, subcortical gray matter structures in the forebrain and brainstem were less severely affected than the cortex. In the cerebellum, seizures appeared to have little effect on the proliferating external granular layer while the closely adjacent internal granular layer showed clear-cut inhibition. The focal nature of protein synthesis inhibition in brain during seizures suggests that the systemic effects of convulsions (e.g. hypoxemia, acidosis or changes in arterial blood pressure) are not a major factor. More likely, protein synthesis inhibition is related to the extent to which various cerebral structures participate in seizure activity.

Amino Acids↗

PubMed1984

Kindling alters the calcium/calmodulin-dependent phosphorylation of synaptic plasma membrane proteins in rat hippocampus.

Septal kindling was associated with an inhibition of the post hoc phosphorylation of several synaptic plasma membrane proteins of rat hippocampus. In control rats, the 32P incorporation into proteins of molecular weights 50,000, 58,000, and 60,000 was markedly stimulated by combined calcium/calmodulin, whereas in kindled animals, the response to combined calcium/calmodulin was reduced. Calcium alone, cAMP, or cGMP modulated 32P incorporation into several synaptic plasma membrane proteins but did not differentiate control from kindled tissues. Both control and kindled rats showed nonspecific inhibition of calcium/calmodulin-stimulated phosphorylation in the post hoc assay by corticotropin and by [Leu]enkephalin. The differences between control and kindled animals were most striking in hippocampus and in the amygdaloid-entorhinal area; less pronounced in cortex, basal ganglia, and brain stem; and not significant in cerebellum, a region where kindling cannot be elicited. An 8-wk period of rest after kindling did not reduce these changes, suggesting that they may be a persistent as the kindling behavior itself.

Animals↗

PubMed1983

Chemical kindling by muscarinic amygdaloid stimulation in the rat.

507 Holtzman rats received injections, through chemitrodes chronically implanted into the basolateral amygdala, of 0.2-1 microliter of sterile isotonic solution containing nanomolar quantities of cholinergic muscarinic agonists and/or antagonists. The bulk of the injected solution diffused only a short distance as judged by autoradiography. Once daily injections of 2.7 nmoles of carbamylcholine, an initially subconvulsive dose, kindled the progressive development of epileptic seizures similar to those seen in electrical amygdaloid kindling. This response was dependent on dose and on interstimulus interval, and once established persisted at least 8 weeks without further stimulation. Spontaneous seizures were observed in some fully kindled animals. No kindling-specific changes were seen by light microscopy. Muscarine (3 nmol) and the active (+), but not the inactive (-), isomer of acetyl-beta-methylcholine also kindled seizures. The action of (+)-acetyl-beta-methylcholine was potentiated by the cholinesterase inhibitor physostigmine. The muscarinic antagonists atropine and quinuclidinyl benzylate (QNB) blocked kindling by carbamylcholine or muscarine. Atropine, QBN and scopolamine greatly reduced agonist-induced seizures in previously kindled rats. Highly significant transfer effects were observed between muscarinic agonists, i.e. muscarine-kindled rats had widespread seizures on their first carbamylcholine exposure and vice versa. Kindled animals had a lowered seizure threshold for muscarinic agonists. Dibutyryl cyclic GMP produced seizures but no kindling. Those results demonstrate that in this model the stimulation of a group of muscarinic cholinergic synapses is both necessary and sufficient to induce a kindled state characterized by both evoked and spontaneous seizures, and support the view that epilepsy can be acquired and expressed transsynaptically.

Amygdala↗