Is the process of localized lysosomal exocytosis responsible for the cytolytic action of killer T-lymphocytes?]

In this paper, we formulate the hypothesis that in the process of target cell lysis a lysosomal enzyme regurgitation, performed by killer cells at the level of the target effector junction, accounts for the target lesion which precedes the lysis (lethal hit). This process of exocytosis, similar to the one described previously in polymorphonuclear neutrophils is supported by cytological studies performed directly on identified killers isolated by micromanipulation. Light and electron microscopy observations confirm a previous report which describes the effector cells rich in lysosomal bodies. In addition, when a killer cell is associated with a target cell to form a conjugate, lysosomes are concentrated near the cell junction and, after incubation at 37 degrees C, acid phosphatases may be detected at the junction. Lysosomal enzyme exocytosis explains why target lysis needs an effector target binding to occur and also the other conditions required for any exocytosis process such as Ca++ in the medium, integrity of the microtubular apparatus, a low level of cyclic AMP and energy dependancy.     

Effect of prostaglandins and dibutyryl cyclic AMP on the morphology of cells in primary astroglial cultures and on metabolic enzymes of GABA and glutamate metabolism

Prostaglandins (PGE1) and dibutyryl cyclic AMP (dBc AMP) induce similar morphological changes in astrocytes obtained in primary cultures. PGE1 and dBc AMP increased 2 enzymes of GABA and glutamate metabolism, GABA-T and AAT, but did not modify GDH and GLN-S. Prostaglandins probably affect the cAMP content of glial cells and act in the same way as dBc AMP on glial cell differentiation.     

Effect of prostaglandins and dibutyryl cyclic AMP on the morphology of cells in primary astroglial cultures and on metabolic enzymes of GABA and glutamate metabolism

Summary Prostaglandins (PGE₁) and dibutyryl cyclic AMP (dBc AMP) induce similar morphological changes in astrocytes obtained in primary cultures. PGE₁ and dBc AMP increased 2 enzymes of GABA and glutamate metabolism, GABA-T and AAT, but did not modify GDH and GLN-S. Prostaglandins probably affect the cAMP content of glial cells and act in the same way as dBc AMP on glial cell differentiation.     

Cyclic AMP is a likely mediator of ovulation in the tsetse fly.

Ovulation in tsetse flies is normally induced by mating, but virgins can be stimulated to ovulate with an injection of dibutyryl cyclic AMP, cholera toxin (a cyclic AMP generator), or aminophylline (a phosphodiesterase inhibitor). Thus, elevation of cyclic AMP is a likely link in the events leading to ovulation.     

Neuronal responses to extracellularly applied cyclic AMP:Role of the adenosine receptor.

At low doses, theophylline blocks the neuronal depressant effects of 5'-AMP, but not cyclic AMP. Higher doses (100 mg/kg) block cyclic AMP responses and reduce the effects of noradrenaline and GABA. It is concluded that cyclic AMP does not depress neurones via the adenosine receptor.     

Role of adenosine A1 and A2 receptors in the regulation of aldosterone production in rat adrenal glands

To investigate the roles of adenosine A1 and A2 receptors in the regulation of aldosterone production, we examined the effects of adenosine and adenosine agonists (N6-cyclohexyl adenosine; selective adenosine A1 receptor agonist and 5'-N-ethylcarboxamine adenosine; selective adenosine A2 receptor agonist) on aldosterone and cyclic AMP production in rat adrenal capsular cells. Neither adenosine nor 5'-N-ethylcarboxamine adenosine caused significant effects on basal aldosterone or cyclic AMP production. Also, adenosine (10(-3) M) showed no consistent effects on aldosterone and cyclic AMP production induced by ACTH. On the other hand, N6-cyclohexyl adenosine exhibited a significant inhibition of basal aldosterone and cyclic AMP production at doses of 10(-4) M and 10(-3) M; furthermore, 10(-3) M N6-cyclohexyl adenosine inhibited aldosterone and cyclic AMP production stimulated by ACTH. These results suggest that adenosine A1 receptors are coupled to and inhibit adenylate cyclase and may be involved in the inhibition of aldosterone production.     

Cyclic AMP is a likely mediator of ovulation in the tsetse fly

Summary Ovulation in tsetse flies is normally induced by mating, but virgins can be stimulated to ovulate with an injection of dibutyryl cyclic AMP, cholera toxin (a cyclic AMP generator), or aminophylline (a phosphodiesterase inhibitor). Thus, elevation of cyclic AMP is a likely link in the events leading to ovulation.We thank J. de Wilde and W.A. Foster for helpful comments. D.L.D.'s research visit to ICIPE was supported by a grant from the Mid-West Universities Consortium for International Activities.     

Effects of TSH and cyclic AMP on the human thyroid cells cultured in a chemically defined medium

Summary In a serum-free, chemically defined medium human thyroid cells elongated remarkably and resembled fibroblastic cells. They retained the cyclic AMP response to TSH and the supplement of medium with TSH or dibutyryl cyclic AMP permitted the preservation of epithelial nature by the cells. Cyclic AMP of the cells of epithelial nature was higher than those of fibroblastic appearance.     

Effects of TSH and cyclic AMP on the human thyroid cells cultured in a chemically defined medium.

In a serum-free, chemically defined medium human thyroid cells elongated remarkably and resembled fibroblastic cells. They retained the cyclic AMP response to TSH and the supplement of medium with TSH or dibutyryl cyclic AMP permitted the preservation of epithelial nature by the cells. Cyclic AMP of the cells of epithelial nature was higher than those of fibroblastic appearance.     

Neuronal responses to extracellularly applied cyclic AMP: Role of the adenosine receptor

Summary At low doses, theophylline blocks the neuronal depressant effects of 5-AMP, but not cyclic AMP. Higher doses (100 mg/kg) block cyclic AMP responses and reduce the effects of noradrenaline and GABA. It is concluded that cyclic AMP does not depress neurones via the adenosine receptor.We are grateful to the Wellcome Trust and Royal Society for travel grants.     

Adenosine promoted accumulation of adenosine 3',5'-monophosphate in rabbit vagus nerve.

Desheathed rabbit vagus nerve has been found to form cyclic AMP when incubated with adenosine. This accumulation of cyclic AMP is inhibited by theophylline but not by antiadrenergic agents, anticholinergic agents or local anaesthetics. Depolarizing media are not able to promote cyclic AMP accumulation in this preparation.     

Beta-adrenergic-stimulated adenylate cyclase activity in normal and EBV-transformed lymphocytes

Beta-adrenergic-associated cyclic AMP accumulation was studied in intact lymphocytes before and after transformation with Epstein-Barr virus into immortal cell lines. Although a marked reduction in isoproterenol-stimulated cyclic AMP synthesis was observed in transformed cells, forskolin-stimulated cyclic AMP accumulation was preserved. A parallel loss of 125-iodocyanopindolol binding sites suggests that the reduction in beta-adrenergic-stimulated AMP synthesis is due to receptor down-regulation.     

Beta-adrenergic-stimulated adenylate cyclase activity in normal and EBV-transformed lymphocytes

Summary Beta-adrenergic-associated cyclic AMP accumulation was studied in intact lymphocytes before and after transformation with Epstein-Barr virus into immortal cell lines. Although a marked reduction in isoproterenol-stimulated cyclic AMP synthesis was observed in transformed cells, forskolin-stimulated cyclic AMP accumulation was preserved. A parallel loss of¹²⁵-iodocyanopindolol binding sites suggests that the reduction in beta-adrenergic-stimulated AMP synthesis is due to receptor down regulation.     

Inhibition of prostaglandin-induced cyclic AMP accumulation in the rat anterior pituitary by alrestatin.

Alrestatin at 25-1 X 10(-4) M inhibited the accumulation of cyclic AMP induced by prostaglandin E2, but not theophylline, in the rat anterior pituitary in vitro. Somatostatin, at lower concentrations, inhibited both; maximal inhibition of the prostaglandin effect was greater with alrestatin. As cyclic AMP is considered to be a mediator in induced-hormonal release, it appears from the present findings that alrestatin may be of potential use in altering hormonal release.     

Analysis of nucleotides bound to the ATP synthetase from spinach chloroplasts isolated under different conditions

Summary The total amount of bound adenine nucleotides in the coupling factor isolated from spinach chloroplasts and its distribution on AMP, ADP and ATP was analyzed after various incubation conditions. During purification of the coupling factor, the distribution of AMP, ADP and ATP is not altered. The coupling factor from deenergized membranes contains approximately 1 ADP, less than 1 ATP, and small amounts of AMP. During phosphorylation the pattern is changed and ATP becomes the dominant species. When exogenous ADP is lacking, phosphate is readily incorporated into ATP. Inhibition of adenylate kinase by AP₅A does not change the distribution pattern of the adenine nucleotides. The distribution pattern shows no integer numbers for the different nucleotides, suggesting that the coupling factor is present in different states in a statistical distribution.Acknowledgment: We thank the Swiss National Foundation for Scientific Research (grant 3.582.79) for generous support.     

Histochemical demonstration of differences in AMP deaminase activity in rat skeletal muscle-fibres

The histochemical method for AMP deaminase confirms that rat slow-twitch fibres contain less AMP deaminase activity than fast-twitch fibres. However, the enzyme appears similarly localized in all fibre types.     

Hormone and forskolin-stimulated cyclic AMP accumulation in human lymphocytes: reliability of longitudinal time measurements

Reliability of measurement of lymphocyte cyclic AMP synthesis in intact cells was estimated by taking 3 successive blood samples during a one-month period from 11 healthy volunteers. Isoproterenol and prostaglandin E1-stimulated cyclic AMP accumulation were used to evaluate the activity of these two receptor activities in human lymphocytes. Forskolin-stimulated cyclic AMP accumulation was used to evaluate the activity of the Ns/catalytic subunit. Only for forskolin was significant reliability observed. For isoproterenol and prostaglandin E1 significant reliability was observed only for male subjects.     

Role of adenosine A1 and A2 receptors in the regulation of aldosterone production in rat adrenal glands

Summary To investigate the roles of adenosine A₁ and A₂ receptors in the regulation of aldosterone production, we examined the effects of adenosine and adenosine agonists (N⁶-cyclohexyl adenosine; selective adenosine A₁ receptor agonist and 5-N-ethylcarboxamine adenosine; selective adenosine A₂ receptor agonist) on aldosterone and cyclic AMP production in rat adrenal capsular cells. Neither adenosine nor 5-N-ethylcarboxamine adenosine caused significant effects on basal aldosterone or cyclic AMP production. Also, adenosine (10^–3M) showed no consistent effects on aldosterone and cyclic AMP production induced by ACTH. On the other hand, N⁶-cyclohexyl adenosine exhibited a significant inhibition of basal aldosterone and cyclic AMP production at doses of 10^–4 M and 10^–3 M; furthermore, 10^–3 M N⁶-cyclohexyl adenosine inhibited aldosterone and cyclic AMP production stimulated by ACTH. These results suggest that adenosine A₁ receptors are coupled to and inhibit adenylate cyclase and may be involved in the inhibition of aldosterone production.     

Opposite effects of beta-adrenoceptor stimulation and 8-bromo-cyclic AMP on potassium efflux in mammalian heart muscle

beta-adrenoceptor stimulation by isoprenaline increases the potassium efflux in beating guinea-pig atria. This effect is not mimicked by 8-bromo-cyclic AMP, a cyclic AMP analogue which exerts a positive inotropic effect in this preparation.     

Histochemical demonstration of differences in AMP deaminase activity in rat skeletal muscle-fibres

Summary The histochemical method for AMP deaminase confirms that rat slow-twitch fibres contain less AMP deaminase activity than fast-twitch fibres. However, the enzyme appears similarly localized in all fibre types.This study was supported by NIH grant AM 21617.