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Biomedical subjects

R D Green

Publications and source records attributed to R D Green.

At least 91 records · Page 5Linked to original sources
PubMed1980

Release of adenosine by C1300 neuroblastoma cells in tissue culture.

Previous work in our laboratory led us to postulate that N2a cells release adenosine into growth medium, where it acts at the extracellular adenosine receptors to modulate the sensitivity of the cells to the cyclic AMP-elevating effect of adenosine [Green, RD, J Pharmacol Exp Ther 201:610, 1977]. We have now devised a high-performance liquid chromatographic (HPLC) procedure capable of quantitating the concentrations of adenosine in cells and tissue culture media. Growth media of N2a cells and a variant of N2a cells deficient in hypoxanthine-guanine phosphoribosyltransferase (HGPRT-) contain 10-20 nM adenosine, while that of a variant deficient in adenosine kinase (AK-) is elevated severalfold. It appears that the concentration of adenosine in growth media is determined by both the rate at which it is released by cells into the medium and the rate at which it is metabolized by adenosine deaminase present in the serum in the growth medium. Both N2a and AK- cells release considerable amounts of adenosine into serum-free medium (SFM) over a short period. Adenosine release is greater from AK- cells and is accelerated by erythro-9-(2-hydroxy-3-nonyl)-adenine (EHNA), a potent adenosine deaminase inhibitor. This accelerated release is retarded by dipyridamole and homocysteine. Surprisingly, dipyridamole and 4-(3-butoxy-4-methoxybenzyl)-2-imidazolidinone (Ro 20 1724), a potent phosphodiesterase inhibitor, stimulate basal adenosine release from N2a but not from AK- cells. It remains to be determined if this is due to an effect of these compounds on adenosine kinase. These results give further support for the hypothesis that adenosine in growth medium modulates the sensitivity of the cells to the cyclic AMP-elevating affect of adenosine, and furthermore they suggest that adenosine in growth media may tonically stimulate adenylate cyclase and affect processes controlled by the cyclic AMP:cyclic AMP-dependent protein kinase system.

Adenosine↗
PubMed1979

Activation of cyclic AMP-dependent protein kinase and stimulation of protein phosphorylation in response to adenosine in C-1300 murine neuroblastoma.

DEAE-cellulose chromatography of the 20,000g supernatant fraction of homogenates of C-1300 murine neuroblastoma (clone N2a) yields one major and two minor peaks of cyclic AMP-dependent protein kinase activity. Assessment of the endogenous activation state of the enzyme(s) reveals that the enzyme is fully activated by the treatment of whole cells with adenosine (10 microM) in the presence of the phosphodiesterase inhibitor Ro 20 1724 (0.7 mM). This treatment produces a large elevation in the cyclic AMP content of the cells. The treatment of whole cells with adenosine alone (1-100 microM) or Ro 20 1724 alone (0.1-0.7 mM) produces minimal elevations in cyclic AMP but nevertheless causes significant activations of cyclic AMP-dependent protein kinase. The autophosphorylation of whole homogenates of treated and untreated cells was studied using [gamma-32P] ATP, sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography. Treatments which activate cyclic AMP-dependent protein kinase selectively stimulate the incorporation of 32P into several proteins. This stimulation is most prominent in the 15,000-dalton protein band. The addition of cyclic AMP to phosphorylation reactions containing homogenate of untreated cells stimulates the phosphorylation of the same protein bands. These results indicate that adenosine may have regulatory functions through its effect on the cyclic AMP:cyclic AMP-dependent protein kinase system.

4-(3-Butoxy-4-methoxybenzyl)-2-imidazolidinone↗
PubMed1979

Pharmacological protection of rabbit kidneys from normothermic ischemia.

This study investigated the influence of the administration of pharmacological agents on the recovery of rabbit kidneys from the effects of 1 hr of situ normothermic ischemia, utilizing acute and chronic models. The agents tested included the diuretics mannitol and furosemide, the vasoactive agents phenoxybenzamine, propranolol, and dopamine, and the membrane stabilizers chlorpromazine and methylprednisolone. A beneficial effect was detected only with the diuretic agents and propranolol when given prior to the ischemic insult.

Animals↗
PubMed1977

Effects of isoproterenol on cyclic AMP and cyclic AMP-dependent protein kinase in developing chick myocardium.

Embryonic chick (7-9 day) and newborn chick myocardia contain one major peak of cyclic AMP-dependent protein kinase activity as assessed by DEAE-cellulose chromatography. Evidence is presented that the cyclic AMP-dependent protein kinase activity ratios (activity in absence of cyclic AMP/activity in presence of added cyclic AMP) of homogenates prepared with low ionic strength buffer reflect the endogenous activation state of the enzyme. The cyclic AMP content of newborn chick myocardium is lower than that of 7--9 day embryonic chick myocardium; the baseline cyclic AMP-dependent protein kinase activity is correspondingly reduced. Isoproterenol produces smaller elevations in cyclic AMP and in the cyclic AMP-dependent protein kinase activity ratio of newborn chick as compared to embryonic chick myocardium. Differences in the ability of isoproterenol to elevate cyclic AMP in the different preparations are not accompanied by appropriate changes in the adenylate cyclase or phosphodiesterase activities of the corresponding broken cell preparations. Studies with the phosphodiesterase inhibitor, Ro 20 1724 indicate that the changes in the ability of isoproterenol to elevate cyclic AMP in the developing chick myocardium are due to changes in the metabolism of the cyclic nucleotide by phosphodiesterase.

4-(3-Butoxy-4-methoxybenzyl)-2-imidazolidinone↗
PubMed1977

Effect of diazoxide, verapamil and compound D600 on isoproterenol and calcium-mediated dose-response relationships in isolated rabbit atrium.

The effect of diazoxide, verapamil and compound D600 on calcium and isoproterenol dose-response relationships was investigated in isolated rabbit atrial preparations. All three agents shifted calcium dose-force relationships parallel and to the right. D600 acted as a competitive antagonist of calcium, as a plot of log (x - 1) vs. -log B resulted in a straight line with a slope of approximately -1.0. Diazoxide, verapamil and D600 also antagonized isoproterenol but in a non-competitive manner. A reduction of calcium in the bathing fluid produced a qualitatively similar non-competitive antagonism of isoproterenol and markedly potentiated diazoxide antagonism of isoproterenol. We conclude that: (1) diazoxide has calcium antagonistic properties similar to the "calcium antagonists" verapamil and D600; (2) these agents act as competitive antagonists of calcium through a specific site that is beyond the beta-adrenergic receptor and in the series of events between the beta-adrenergic receptors and inotropic response in myocardial tissue.

Adrenergic beta-Antagonists↗
PubMed1977

Sensitivity of the developing chick myocardium to the positive inotropic effects of calcium and isoproterenol.

The present results show that the sensitivity of the chick myocardium to the positive inotropic effect of Ca++ decreases during development and that the Ca++ concentration of the physiological solution used must be lowered below 'normal' to study the effects of positive inotropic agents in preparations from younger embryos. Isoproterenol elicits positive inotropic responses in 7-9 day embryonic ventricle and in newborn chick atria; however, the 4-day embryonic myocardium is unresponsive to isoproterenol.

Aging↗
PubMed1977

Control of cyclic adenosine 3':5'-monophosphate-elevating effect of adenosine in C-1300 murine neuroblastoma in tissue culture.

The elevation of cyclic adenosine 3':5'-monophosphate (cyclic AMP) in response to adenosine in C-1300 murine neuroblastoma (clone N2a) in surface culture is increased in magnitude in cultures pretreated overnight with theophylline or adenosine deaminase. This "potentiating" effect of theophylline takes time to develop and is blocked by cycloheximide. The cyclic AMP-elevating effect of adenosine decreases in magnitude as the cultures approach confluence. This reduced responsiveness is reversed by the overnight treatment with theophylline. It is hypothesized that adenosine is continually released by the cells to the growth medium and that this adenosine acts extracellularly to modulate the sensitivity of the cells to the cyclic AMP-elevating effect of adenosine.

4-(3-Butoxy-4-methoxybenzyl)-2-imidazolidinone↗
PubMed1975

The effects of chronic ganglion blockade and chronic cholinesterase inhibition on the sensitivity of rabbit stomach muscularis to cholinergic and adrenergic agonists.

The EC50 values, KA values and efficacies (e) of selected agonists have been determined in strips of stomach muscularis from control, chlorisondamine-pretreated and O, O-diethyl S[2(ethylthio)ethyl]phosphodithioate (disulfoton)-pretreated rabbits. Strips from chlorisondamine-pretreated animals were supersensitive to carbachol but normosensitive to phenylephrine; the e but the KA of carbachol was affected by this treatment. The KB of atropine was unchanged. Strips from animals treated with disulfoton were subsensitive to phenylephrine but normosensitive to amidephrine and carbachol; both the e and KA of phenylephrine appeared to be elevated by this pretreatment. The KB of phentolamine remained unchanged. These results indicate that the chlorisondamine-induced supersensitivity is the result of a change beyond the level of the cholinergic receptors. In contradistinction, the subsensitivity to phenylephrine after disulfoton pretreatment may result at least in part, from a qualitative change in the alpha adrenergic receptors in the stomach muscularis.

Acetylcholinesterase↗