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Biomedical subjects

Y H Ehrlich

Publications and source records attributed to Y H Ehrlich.

At least 55 records · Page 3Linked to original sources
PubMed1986

Proteolysis of the platelet surface: dissociation of shape change from aggregation.

Stimulation of intact platelets by ADP results in a shape change followed by aggregation in the presence of fibrinogen. ADP was found to induce a shape change in chymotrypsin-treated platelets that was similar in extent and initial velocity to that of intact (untreated) platelets. Scanning-electron microscopy verified an ADP-induced shape change in chymotrypsin-treated platelets. This shape change could be completely blocked by stimulators of platelet adenylate cyclase (forskolin, prostaglandin E1, and prostacyclin). On the other hand, the aggregation of chymotrypsin-treated platelets by fibrinogen was not dependent on the presence of ADP and could not be blocked by forskolin, prostaglandin E1, or prostacyclin, even though the levels of cyclic AMP (cAMP) formed in chymotrypsin-treated platelets were comparable to levels that completely inhibited the ADP-induced aggregation of intact platelets. This lack of inhibition of platelet aggregation was not due to degradation of the adenylate cyclase or prostaglandin receptors, since chymotrypsin-treated platelets were found to have a functional adenylate cyclase system that could be stimulated by forskolin, prostaglandin E1, and prostacyclin and inhibited by ADP and epinephrine, similar to that of intact platelets. These results provide direct evidence that cAMP does not interact with fibrinogen binding sites once they have become permanently exposed on the surface of platelets. Pretreatment of platelets with chymotrypsin therefore appears to be a useful tool that allows for the dissociation of platelet shape change from aggregation, without inhibiting either response.

Adenosine Diphosphate↗
PubMed1986

Exposure of fibrinogen receptors in human platelets by surface proteolysis with elastase.

Human platelets that were preincubated with porcine elastase aggregated spontaneously upon the addition of fibrinogen. Maximal aggregation to fibrinogen was observed with platelets pretreated with an elastase concentration of 111 micrograms/ml, and half-maximal aggregation occurred after treatment with 11 micrograms/ml elastase. Binding of radiolabeled fibrinogen to elastase-treated platelets was specific, saturable, and showed a single class of 48,400 +/- 9,697 fibrinogen-binding sites per platelet with a dissociation constant of 6.30 +/- 1.48 X 10(-7) M. ATP, apyrase, and the stimulators of platelet adenylate cyclase forskolin, prostaglandin E1, prostacyclin, and N6, 2'-O-dibutyryl cyclic AMP did not inhibit the fibrinogen-induced aggregation of elastase-treated platelets. EDTA completely blocked the initiation of aggregation and reversed the fibrinogen-induced aggregation of elastase-treated platelets. Monoclonal and polyclonal antibodies directed against glycoproteins (GP) IIb and IIIa completely blocked the fibrinogen-induced aggregation of elastase-treated platelets. Immunoprecipitates with these antibodies obtained from detergent extracts of surface-radiolabeled, intact, and elastase-treated platelets contained the glycoproteins IIb and IIIa. We conclude that surface proteolysis by low concentrations of elastase can expose fibrinogen-binding sites associated with GPIIb and GPIIIa on the platelet surface, resulting in spontaneous aggregation upon the addition of fibrinogen. These findings may be relevant to hemostatic changes observed in patients with increased levels of circulating elastase.

Adenosine Triphosphate↗
PubMed1985

Alpha 2-adrenergic receptor-mediated regulation of adenylate cyclase in the intact human platelet. Evidence for a receptor reserve.

The alpha 2-adrenergic receptor on the human platelet is known to mediate the inhibition of adenylate cyclase activity. A comparison of the binding and response properties of intact cells revealed that the full agonists norepinephrine and epinephrine inhibit cyclic AMP accumulation with apparently higher affinity than they exhibit in inhibiting the binding of [3H]yohimbine. Additionally, Hill coefficients of the occupancy curves of the agonists were less than unity, suggesting the presence of a heterogeneous receptor population in intact platelets under conditions that permit robust inhibition of cyclic AMP accumulation. The partial agonist clonidine was found to possess the same affinity in the binding assay as in the response assay. These data are consistent with the presence of a receptor reserve in this system, a suggestion that was confirmed in experiments utilizing the irreversible alpha 2 antagonist phenoxybenzamine. The IC50 (100 nM) derived from the blockade of [3H]yohimbine binding by phenoxybenzamine was significantly less than the IC50 (550 nM) for the corresponding reversal by phenoxybenzamine of the effects of norepinephrine on cyclic AMP accumulation. Further studies demonstrated a rightward shift in the dose-response curves for the inhibition by norepinephrine of cyclic AMP accumulation following pretreatment with increasing phenoxybenzamine concentration. These data consistently indicated that occupancy of approximately 10% of the alpha 2-adrenergic receptors by norepinephrine elicits a half-maximal adenylate cyclase response. The relationship of these findings to current models of receptor-effector coupling is discussed.

Adenylyl Cyclases↗
PubMed1984

Platelet-activating factor-induced aggregation of human platelets specifically inhibited by triazolobenzodiazepines.

Platelet-activating factor (PAF), a naturally occurring phospholipid, is a potent activator of various biological processes, including platelet aggregation. The mechanisms by which PAF acts are largely unknown, partly because of the lack of specific inhibitors for PAF-elicited responses. It was found that in washed human platelets the psychotropic triazolobenzodiazepine drugs alprazolam and triazolam potently inhibited PAF-induced changes in shape, aggregation, and secretion. The effects were specific for PAF activation, since the responses of human platelets to adenosine diphosphate, thrombin, epinephrine, collagen, arachidonate, and the calcium ionophore A23187 were not inhibited by the triazolobenzodiazepines. These psychotropic drugs should be useful in investigating the possibility that PAF or PAF-like phospholipids play a role in neuronal function and in elucidating biochemical mechanisms activated specifically by PAF in a variety of cells.

Adenosine Diphosphate↗
PubMed1984

Activation of adenylyl cyclase by preincubation of rat cerebral-cortical membranes under phosphorylating conditions: role of ATP, GTP, and divalent cations.

The effects of preincubation under phosphorylating conditions on adenylyl cyclase activity were studied in preparations containing synaptic membranes from rat cerebral cortex. Preincubation of the membranes with 2 mM ATP and 10 mM MgCl2 resulted in a 50% increase of adenylyl cyclase activity which withstood sedimentation and washing. This activation was maximal after 5 min of preincubation, was reversed after longer preincubations, and paralleled the time course of endogenous phosphorylation-dephosphorylation of proteins observed under these conditions. The activation showed a critical requirement for Mg2+ ions and was dependent on ATP concentration. Similar activation was observed after preincubation of cerebral-cortical membranes with adenosine-5'-O-(3-thiophosphate) (ATP gamma S), but this activation was not reversed by prolonged preincubation times. The activation by ATP gamma S was potentiated severalfold by including synaptoplasm in the preincubation. Further experiments indicated that the activity of nucleoside diphosphokinase, which converts ATP gamma S to guanosine-5'-O-(3-thiophosphate) (GTP gamma S), could account for this potentiation. Preincubation of washed membranes for 5 min with 10 microM GTP and 10 mM MgCl2 also produced a 50% activation of adenylyl cyclase which withstood sedimentation and washing and was reversed by longer preincubations. Endogenous phosphorylation of specific protein components in the membranes during the preincubation was examined by including radioactively labeled nucleoside thiophosphates in the preincubation medium. Incorporation of 35S from [35S]ATP gamma S into a protein component with apparent Mr of 54,000 daltons ( 54K ) correlated significantly with the activation of adenylyl cyclase by ATP gamma S. Thiophosphorylation of the 54K protein was potentiated by addition of GDP to reactions carried out with [35S]ATP gamma S. Endogenous activity utilizing [gamma-32P]GTP as a phosphate donor also preferentially phosphorylated the 54K protein band. These results support previous suggestions that protein phosphorylation plays a role in the regulation of adenylyl cyclase activity. Among the numerous membrane-bound phosphoproteins in rat brain, we have identified a specific protein component with an apparent Mr of 54,000 daltons as the most likely candidate for involvement in this mode of regulation. This 54K protein, which is a principal substrate for a GTP-preferring protein kinase activity in brain membranes, can now be at the focus of investigations attempting to demonstrate a direct role for protein phosphorylation in adenylyl cyclase regulation.

Adenosine Triphosphate↗
PubMed1983

Automated analysis of stereotypic behavior induced by psychomotor stimulants.

A newly developed rotation sensing device has been applied to the continuous monitoring of animal movement. Animals treated with morphine, amphetamine or apomorphine display different stereotypic movements which can be distinguished by the apparatus. Initial studies have indicated that the apparatus is able not only to identify but also to quantitate some measures of stereotypic behavior. For example, the number and direction of rotations (a measure of motor asymmetry), frequency of changes in movement direction (a measure of stereotypic movement) and periods of cessation of movement are affected differentially with acute morphine, apomorphine or amphetamine treatment. Moreover, using this apparatus, morphine was shown to increase the degree of rotational asymmetry of normal animals and of animals with unilateral lesions of the nigrostriatal pathway.

Amphetamine↗
PubMed1982

A device for the automated monitoring of stereotypic behavior.

1. A novel rotation sensing device has been developed which consists of a linear potentiometer attached to the top of a spherical testing chamber. The potentiometer constantly monitors the precise angular position of the animal inside the chamber. 2. This device allows the automated monitoring of direction and rate of rotation, as well as a variety of other stereotypic behaviors. 3. The effects of morphine, apomorphine and amphetamine on motor activity were assessed by the apparatus.

Amphetamine↗
PubMed1981

Protein phosphorylation mediates effects of isoproterenol on adenylate cyclase activity in rat cortical membranes.

The effects of (-)-isoproterenol on adenylate cyclase activity were studied in rat cerebral cortical membranes prepared and assayed in the presence of calcium ions. In assays carried out in the presence of high Mg2+ concentrations (5-10 mM) and of Ca2+ in the micromolar range, addition of 1-100 micro M (-)-isoproterenol caused over 50% inhibition of adenylate cyclase activity. Since these conditions are optimal for supporting endogenous phosphorylative activity in synaptic membranes, we tested whether the observed effects are mediated by changes in the phosphorylation of specific proteins in these membranes. This was done by preincubation of lysed synaptosomes under phosphorylating conditions in the presence and absence of isoproterenol followed by extensive washes and analysis of cyclic AMP formation in resuspended membranes. Addition of (-)-isoproterenol to the preincubation resulted in a 30% decrease of adenylate cyclase activity in the reincubation. Inclusion of [gamma-32P]ATP in the preincubation and examination of the phosphorylation state of specific proteins in membranes entering the reincubation revealed that (-)-isoproterenol inhibited the phosphorylation of a specific protein band with apparent molecular weight of 47,000 (designated band F). These results support the hypothesis that alterations in membrane protein phosphorylation induced by neurotransmitters play a role in the regulation of adenylate cyclase activity.

Adenosine Triphosphate↗
PubMed1979

Endorphins, dopamine, and schizophrenia.

The theory that alterations of dopaminergic synaptic transmission may play a role in the pathogenesis of schizophrenia is widely accepted. A more recent theory links the endorphin system to the etiology of schizophrenia. We propose that these two theories may be combined into a single model. Recent neurochemical and pharmacological findings have indicated close functional relationships between the endorphin and dopamine systems. Endorphins modulate dopaminergic synaptic transmission by exerting both presynaptic and postsynaptic effects. On the molecular level, this modulation may involve the activity of nucleotide cyclases and protein phosphorylation systems. Thus, the dopaminergic neuronal hyperactivity, currently believed to be related to schizophrenia, may be caused by a primary alteration in the endorphin system. Several hypotheses about the nature of that alteration have been advanced and tested in therapeutic experiments with schizophrenic patients. These experiments have not yet yielded definitive results.

Antipsychotic Agents↗