Effects of UVB irradiation on epidermal adenylate cyclase responses in vitro: its relation to sunburn cell formation.

UVB irradiation augmented the beta-adrenergic adenylate cyclase response of pig skin epidermis in vitro. The effect was observed 2-4 h following the irradiation and lasted at least for 48 h. There was no significant difference in cyclic AMP phosphodiesterase activity between control and UVB-irradiated epidermis at lower irradiation dose (150 mJ/cm2), which is the dose of the most marked beta-adrenergic augmentation effect. The augmentation effect was specific to the beta-adrenergic system; adenosine and histamine adenylate cyclase responses were unchanged or decreased depending on the irradiation dose. Histologically, marked sunburn-cell formation was observed following the UVB irradiation. It has been suggested that oxygen intermediates generated by ultraviolet radiation participate in sunburn-cell formation. The addition of superoxide dismutase (SOD) in the incubation medium significantly inhibited sunburn-cell formation. On the other hand, the beta-adrenergic augmentation effect was not affected by the addition of SOD. Other scavengers of oxygen intermediates (catalase, catalase + SOD, xanthine, or mannitol) did not inhibit the UVB-induced beta-adrenergic augmentation effect. Further, superoxide-anion generating systems (hypoxanthine-xanthine oxidase system and acetaldehyde-xanthine oxidase system) revealed no stimulatory effect on the beta-adrenergic response of epidermis. These results indicate that (a) the UVB-induced beta-adrenergic augmentation effect is inherent to skin and does not depend on systemic factors such as inflammatory infiltrates following UVB irradiation; (b) in contrast to sunburn-cell formation, induction of the beta-adrenergic adenylate cyclase response is not directly associated with oxygen intermediates generated by UVB irradiation.

Denervation supersensitivity of refractoriness in noninfarcted areas apical to transmural myocardial infarction.

Denervation supersensitivity was demonstrated in anesthetized dogs 5 to 10 days after transmural myocardial infarction produced by latex embolization of a diagonal branch of the left anterior descending coronary artery. Sympathetic efferent denervation in noninfarcted myocardium apical to the infarction was demonstrated by a 90% depletion of myocardial norepinephrine content in the apical (45 +/- 15 pg norepinephrine/g tissue) vs basal (437 +/- 76 pg/g tissue) regions and by the lack of effective refractory period (ERP) shortening during bilateral ansae subclaviae stimulation in 34% of sites apical to the infarction. Supersensitivity in the area apical to the infarction was manifested by an exaggerated shortening of the ERP during both norepinephrine and isoproterenol infusions, with an upward and leftward shift in the dose-response curves in the apical vs basal regions (p less than .001). The cellular mechanism for denervation supersensitivity did not involve detectable changes in the beta-adrenergic receptor adenylate cyclase system. There was no difference in the density of beta-adrenergic receptors ([125I]-cyanopindolol) in the apical (268.6 +/- 22.7 fmol/mg protein) vs the basal (253.5 +/- 24.8 fmol/mg protein) regions. Adenylate cyclase activity stimulated by guanosine triphosphate plus isoproterenol was slightly greater in the apical (58.7 +/- 17.4%) than in the basal (49.6 +/- 10.9%) region, but this difference did not reach statistical significance (p = .068). Muscarinic modulation of beta-receptor coupling (oxotremorine attenuation of guanosine triphosphate plus isoproterenol-stimulated adenylate cyclase activity) also was not significantly different at the apical (31.6 +/- 17.5% inhibition) and basal (21.4 +/- 20.9% inhibition) sites. These data show that a transmural myocardial infarction produces denervation supersensitivity in areas apical to the infarction, but in this preparation no differences in the total number or a redistribution of beta-adrenergic receptors or adenylate cyclase activity were detected.

[Mechanisms of the disorder of the hormonal regulation of plasma membrane adenylate cyclase in the rat liver during early ontogeny under in vitro irradiation]. / Mekhanizmy narusheniia gormonal'noi reguliatsii adenilattsiklazy plazmaticheskikh membran pecheni krysy v khode rannego ontogeneza v usloviiakh oblucheniia in vitro.

Possible reasons for the previously detected inhibitory effect of gamma-radiation (100-1000 Gy in vitro) on isoproterenol-stimulated activity of adenylate cyclase (AC) of the liver plasma membranes of a 20-day-old rat fetus were investigated. It was discovered that the number of beta-adrenergic receptors was reduced. Analysis of basal and fluorine-stimulated activities of AC attests to the radiation-induced damage to catalytic and N-regulatory proteins. The radiation-induced derangement of membrane lipids, where AC functions, is shown.

Demonstration of beta-2 adrenergic receptors of high coupling efficiency in human neutrophil sonicates.

Using highly purified (greater than 95%) neutrophil (PMN) sonicates, we have studied the beta adrenergic (beta 1/beta 2) pattern of adenylate cyclase activation following agonist stimulation and have assessed the coupling characteristics of the beta adrenergic receptor to the adenylate cyclase enzyme. Adenylate cyclase was highly responsive to agonist activation, with peak isoproterenol (100 microM) stimulation resulting in the generation of 119 +/- 9.5 (mean +/- S.E.M.) pmol/mg/min cyclic AMP (224% above basal levels) compared to 171.7 +/- 8.6 following NaF (10 mM) stimulation. The agonist pattern of adenylate cyclase activation suggested the presence of beta-2 adrenergic receptors, since isoproterenol with a Kact of 0.7 microM was more potent than epinephrine (Kact = 8.5 microM) or norepinephrine (Kact = 90 microM). Butoxamine (beta-2 antagonist) was approximately 25 times more potent than practolol (beta-1 antagonist), with KDs of 0.75 microM and 17.5 microM, respectively. Receptor coupling efficiency was determined by measuring isoproterenol binding and adenylate cyclase activation with the same PMN sonicates and incubation conditions for each assay. The apparent KD for isoproterenol binding was 2.82 +/- 0.53 muM, the Kact was 0.47 +/- 0.05 muM, and the mean KD/Kact ratio was 6.5. Highly coupled receptor-enzyme systems have ratios greater than or equal to 1. Using sucrose gradient-purified PMN sonicates, we found that isoproterenol required the guanine nucleotides GTP or Gpp(NH)p to activate adenylate cyclase. The GTP effect on adenylate cyclase responsivenss to isoproterenol was associated with a 10-fold decrease in the isoproterenol binding affinity. These studies suggest that the human PMN has beta-2 adrenergic receptors which are highly coupled to the adenylate cyclase enzyme. We believe that this tissue offers a suitable model for the study of beta-2 adrenergic receptors and the mechanism of hormone-induced adenylate cyclase activation in man.

Adenylate cyclase in leukocytes from patients with cystic fibrosis.

Lymphocytes from patients with cystic fibrosis produce significantly less cAMP in response to beta-adrenergic stimulation than do cells from healthy persons (p less than 0.0005) or patients with bronchiectasis (p less than 0.005). Adenylate cyclase in the basal state or stimulated by GMPPNP or PGE1 is normal, but isoproterenol-stimulated adenylate cyclase activity is significantly (p less than 0.005) reduced in lymphocyte membrane preparations from patients with cystic fibrosis. Granulocytes from patients with cystic fibrosis also produce significantly less cAMP in response to beta-adrenergic stimulation than do cells from healthy subjects (p less than 0.0005) or subjects with bronchiectasis (p less than 0.05). Adenylate cyclase activity in granulocyte homogenates from patients with cystic fibrosis is normal in the basal state or when stimulated by GMPPNP or PGE1 but is significantly (p less than 0.05) reduced compared to normal when stimulated by isoproterenol. These data suggest that the defect is not related to adenylate cyclase itself or to availability of substrate or cofactor to adenylate cyclase, and they may be indicative of a more generalized membrane defect in cystic fibrosis.