Glial fibrillary acidic protein and Alzheimer's disease.

A major protein associated with Alzheimer's disease (AD) was detected by an electrophoretic study of temporal cortex obtained at autopsy from patients affected with AD, non-AD dementia, and normal controls matched for age and sex. A markedly increased amount of a 50,000 dalton molecular weight protein, which has been identified as glial fibrillary acidic protein (GFAP), was observed in the crude nuclear fraction of temporal cortex from AD patients. These electrophoretic data may reflect the presence of GFAP immunopositive astrocytic processes that have been shown by immunocytologic methods to infiltrate the neurofibrillary tangles that characterize AD.

Relationship of membrane physical properties to alcohol dependence in mice selected for genetic differences in alcohol withdrawal.

Genetic selection based on severity of withdrawal seizures following inhalation of ethanol vapor has produced two lines of mice, WSR (withdrawal seizure resistant) and WSP (withdrawal seizure prone), that differ markedly in withdrawal signs. In the present study, we report that these mice also differed in the severity of withdrawal seizures following consumption of an ethanol-containing liquid diet but did not differ in ethanol intake. In contrast to ethanol withdrawal seizures, the lines displayed similar sensitivity to electrical- or pentylenetetrazole-induced seizures. These results suggest that the lines differ in the development of physical dependence on ethanol rather than seizure sensitivity per se. Because decreased synaptic membrane fluidity has been associated with ethanol dependence, we used fluorescence polarization of diphenylhexatriene and trimethylammonium-diphenylhexatriene to evaluate membrane fluidity in WSP and WSR mice fed lab chow, an ethanol-containing liquid diet, or an isocaloric sucrose-containing liquid diet. Fluidity of brain synaptic membranes was identical for WSP and WSR mice fed lab chow. The control liquid diet did not alter membrane fluidity, and the ethanol diet decreased fluidity equally for WSP and WSR mice. Thus, the genetic difference in development of ethanol dependence found in these lines was not reflected in the physical properties of brain membranes.

Specific ethanol withdrawal seizures in genetically selected mice.

We are selectively breeding mice prone (WSP) and resistant (WSR) to ethanol withdrawal seizures assessed by handling induced convulsions (HIC). The possibility that differences between the lines in HIC scores are a result of differences in general CNS excitability not specific to ethanol withdrawal was examined. Using treatments which produce generalized seizures (electroconvulsive shock, strychnine, and flurothyl) and gamma amino-butyric acid (GABA) antagonists (picrotoxin, bicuculline, and pentylentetrazol), the ED50 for seizures was determined in the selected lines. In addition, the sensitivity of WSP and WSR mice to the anticonvulsant actions of ethanol against each treatment was determined. Neither the convulsant amperage 50 (CA50) for ECS nor the ED50 for any drug treatment differed for the selected lines. When ethanol (1.5 g/kg) was administered prior to ECS, there was a dramatic differential suppression of ECS in the lines: the CA50 of WSR mice was elevated 5-fold, whereas the CA50 of WSP mice increased only two fold. Ethanol pretreatment also elevated the ED50 for strychnine and flurothyl in WSR mice significantly more than WSP mice, but the line difference was smaller than for the anticonvulsant effect against ECS. The ED50s for the GABA antagonists were not different between the WSR and WSP lines after ethanol pretreatment. We conclude that genetic selection is producing lines of mice that differ specifically in the degree of seizure severity caused by withdrawal from ethanol physical dependence and not in generalized CNS excitability. An increased sensitivity to the anticonvulsant effects of ethanol against some convulsant treatments has appeared as a correlated response to selection in the WSR line.

Chromosomal assignment of the gene for the human beta 2-adrenergic receptor.

Chinese hamster fibroblasts (CHW) fail to physiologically respond to the beta-adrenergic agonist isoproterenol with an increase in cellular cAMP. This unresponsiveness is due to a lack of beta-adrenergic receptors as indicated by an absence of specific binding of 125I-labeled hydroxybenzylpindolol (125I-HYP) to CHW plasma membranes. Preparation of somatic cell hybrids between CHW and human peripheral blood leukocytes led to the selection of a panel of 15 human-Chinese hamster cell hybrid clones, some of which had beta 2-adrenergic receptors (specifically bound 125I-HYP) and responded to isoproterenol (accumulated cAMP). Biochemical analysis of independent cloned cell hybrids indicated that beta 2-adrenergic receptor density and the intensity of the associated physiological response were closely correlated (r = 0.98). Both of these parameters were concordant in all cell hybrids with the presence of human chromosome 5. All other human chromosomes could be ruled out. These results suggest that the structural gene for the beta 2-adrenergic receptor is found on human chromosome 5.

Assessment of red cell sodium transport in essential hypertension.

Abnormal erythrocyte Na+ transport has been reported in patients with essential hypertension and some first-degree relatives. The two major techniques now employed for estimating Na+ transport--Na+/Li+ countertransport and Na+/K+ cotransport--are rather intricate and time consuming. Furthermore, the precise nature of the transport processes being measured is not clear. We have developed a simpler, more direct technique based on measurement of 22Na+ accumulation by erythrocytes. 22Na+ uptake by red cells from patients with essential hypertension averages twice normal. Indeed, of 21 patients with essential hypertension, only 2 patients had values within the upper end of the normal range. In 12 patients with secondary hypertension and no family history of essential hypertension, erythrocyte 22Na+ accumulation was within normal limits. Control experiments indicate that our technique for estimating red cell 22Na+ uptake is highly reproducible and shows little day-to-day variation. This procedure for the assessment of erythrocyte Na+ transport should be useful in differential diagnosis and the presymptomatic identification of individuals genetically prone to essential hypertension.

The adrenergic responsiveness of Down syndrome cells.

Platelets and cultured fibroblasts were used to examine the hormonal response of normal diploid and Down syndrome (trisomy 21) cells. An enhanced physiological response to catecholamine hormones was found in the trisomy 21 cells. This effect was specific for the adrenergic agonists since other hormones showed equivalent responses in the normal diploid and trisomy 21 cell types. Radioligand analysis of the alpha-adrenergic receptors in trisomy 21 platelets indicated no significant difference in receptor density or ligand affinity. Enzymatic analysis of the platelet adenylate cyclase activity in vitro showed no major changes in catecholamine sensitivity between the normal and Down syndrome preparations. Somatic cell genetic studies using human-hamster hybrids indicated that a gene coding for the beta-adrenergic receptor was not located on chromosome 21 but on human chromosome 5. These receptor, enzymic, and somatic cell genetic studies argue against a gene-dosage effect involving either the adrenergic hormone receptors or a component of the adenylate cyclase enzyme in the trisomy 21 cells. Identifying the gene(s) on chromosome 21 and the molecular mechanism(s) responsible for the exaggerated response of Down syndrome cells to catecholamines are the current objectives of this research program.

Postmortem stability of dopamine-sensitive adenylate cyclase, guanylate cyclase, ATPase, and GTPase in rat striatum.

The stability of dopamine-sensitive adenylate cyclase, guanylate cyclase, ATPase, and GTPase was measured in homogenates of rat striatal tissue frozen from 0 to 24 h postmortem. ATPase, GTPase, and Mg2+-dependent guanylate cyclase activities showed no significant change over this period. Mn2+-dependent guanylate cyclase activity was stable for 10 h postmortem. Basal and dopamine-stimulated adenylate cyclase activity decreased markedly during the first 5 h. However, when measured in washed membrane preparations, these adenylate cyclase activities remained stable for at least 10 h. Therefore, the postmortem loss of a soluble activator, such as GTP, may decrease the adenylate cyclase activity in homogenates. These results are not consistent with an earlier suggestion that there is a postmortem degradation of the enzyme itself. Other kinetic parameters of dopamine-sensitive adenylate cyclase can also be measured independently of postmortem changes. Thus, it is possible to investigate kinetic parameters of dopamine-sensitive adenylate cyclase, guanylate cyclase, ATPase, and GTPase in human brain obtained postmortem.

Down syndrome fibroblasts are hyperresponsive to beta-adrenergic stimulation.

The hormonal response to human skin fibroblasts after exposure to beta-adrenergic agonists, prostaglandin E1 (PGE1), and cholera toxin was monitored by intracellular cyclic AMP accumulation. Down syndrome (DS; trisomy 21) cells had an approximately 10-fold greater response to beta-adrenergic agonists than did either normal diploid skin fibroblasts or other aneusomic fibroblast strains (trisomy 13, 18, and 22). The altered response in DS fibroblasts was specific for beta-adrenergic agonists, because treatment of DS or control cells with PGE1 or cholera toxin resulted in the same degree of cyclic AMP accumulation. Experiments with 3-isobutyl-1-methylxanthine, a cyclic nucleotide phosphodiesterase inhibitor, indicated that the increased response of DS fibroblasts was not primarily a function of altered cyclic AMP degradation. Monosomy 21 cells responded less than normal diploid fibroblasts to stimulation by the beta-adrenergic agonist isoproterenol. These findings suggest that genetic information on chromosome 21 participates in regulating the beta-adrenergic response of human fibroblasts.