Sodium-dependent accumulation of 5-hydroxytryptamine by rat blood platelets.

1. The influence of sodium and potassium on the accumulation of 5-hydroxytryptamine (5-HT) by rat blood platelets was investigated.2. An absolute dependence of 5-HT uptake on the sodium concentration in the medium was found.3. Removal of potassium reduced the uptake by about 60%. High concentrations of potassium inhibited sodium-dependent accumulation.4. The observations have been discussed in terms of a carrier-mediated transport process for 5-HT operating in the platelet membrane.

Relationship between internal Na + -K + and the accumulation of 14 C-5-hydroxytryptamine by rat platelets.

1. 5-Hydroxytryptamine (5-HT) transport has been investigated in rat blood platelets poisoned with dinitrophenol-sodium fluoride or ouabain.2. The inhibition of transport produced by different concentrations of the metabolic inhibitors has been correlated with changes in the internal Na(+) and K(+) concentrations of the platelets.3. Platelets poisoned in a high K(+) medium maintained a high internal K(+) concentration in the absence of cellular metabolism. When transferred to Krebs solutions containing different concentrations of Na(+) they accumulated 5-HT by a process that was related to the magnitudes of the internal and external Na(+) concentrations.4. The results are consistent with the hypothesis that the spontaneous movement of ions through the platelet membrane is capable of providing, at least in part, the energy requirements for 5-HT transport.

The influence of metyrapone on the synthesis and release of prostaglandins from the pregnant rat uterus in vitro.

1 Metyrapone (150 mg/kg, s.c. or i.p.) an inhibitor of corticosteroid biosynthesis, significantly reduced the release of prostaglandins of the F-type from isolated preparations of pregnant rat uteri in vitro, on day 22 - the expected day of delivery. 2 Metyrapone and indomethacin administered in vitro both inhibited the conversion of 14C-arachidonic acid to prostaglandin E2 by homogenates of day 22 pregnant rat uteri. Metyrapone was approximately 150 times less potent than indomethacin. Although indomethacin also inhibited prostaglandin F2alpha production, metyrapone stimulated synthesis of this prostaglandin. The differential inhibition of prostaglandin synthesis by metyrapone may reflect sensitivity of the inhibitor to changes in experimental conditions. 3 Inhibition of prostaglandin synthesis may explain the effects of metyrapone on parturition in the rat.

Effect of selective proteolysis on the accumulation of 5-hydroxytryptamine by intact rat blood platelets.

1 The effect of perturbation of intact blood platelets with proteolytic enzymes was studied with respect to 5-hydroxytryptamine (5-HT) transport, adenine transport and intracellular Na(+) and K(+) levels.2 Leucine aminopeptidase and thrombin reduced 5-HT transport, released 5-HT from pre-labelled platelets and disturbed the gradient to monovalent cations. Leucine amino-peptidase, but not thrombin, inhibited adenine transport.3 alpha-Chymotrypsin and carboxypeptidases A and B were without effect on the parameters studied.4 Trypsin selectively reduced 5-HT transport. It did not release 5-HT from blood platelets or inhibit adenine transport.5 The action of proteolytic enzymes is discussed in terms of proteins localized in the external membrane that may function in part as membrane carriers.

Superoxide anions enhance platelet adhesion and aggregation.

1. Superoxide dismutase (SOD, 60 u ml-1) or ferricytochrome c (70 microM) significantly inhibited thrombin-stimulated platelet adhesion to gelatin-coated plastic, whereas catalase (1000 u ml-1) or mannitol (1 mM) had no effect. 2. The platelet aggregation induced by low concentrations of thrombin (causing less than 45% maximal change in light transmission) was inhibited by SOD. Catalase or mannitol had no effect on platelet aggregation. 3. Pyrogallol (an O2- generator) enhanced both platelet adhesion to gelatin-coated plastic and platelet aggregation induced by thrombin; this enhancement was neutralized by SOD. 4. These results indicate that O2- increase both platelet adhesion and aggregation, whereas other free radicals such as hydrogen peroxide or hydroxyl radicals are not involved.

Evidence for a possible foetal control of prostaglandin release from the pregnant rat uterus in vitro.

The release of prostaglandin-like material and these spontaneous contractions of individual horns from the pregnant rat uterus in vitro have been studied on day 22 of pregnancy - the expected day of delivery. Removal of foetuses (retaining placentae in utero) from one or both uterine horns on day 16 or 17 significantly reduced prostaglandin F release and spontaneous activity. Rats which had been made unilaterally pregnant after ligation of one uterine horn, exhibited a decrease in prostaglandin F output from both horns. Uterine activity and prostaglandin release were increased in quiescent uteri by the addition of arachidonic acid (5 mug/ml) or phospholipase A (160 mu./ml); the effects were abolished by indomethacin (20 mug/ml). However, the stimulation of uterine activity by PGF2alpha (30-60 ng/ml) was not affected by indomethacin. It is concluded that the release of prostaglandins from the pregnant rat uterus in vitro at term is related to the presence of viable foetuses.

The influence of pH on aggregation of human washed platelets induced by thrombin or collagen.

The anti-aggregating activity of L-arginine and its analogues has been investigated in washed human platelets. The ability of these compounds to inhibit platelet aggregation induced by thrombin or collagen was mimicked by a change in the external pH of the buffer from pH 8.0 to pH 7.4. Of the several analogues tested, only benzoyl-L-arginine ethylester (BAEE) inhibited thrombin- but not collagen-induced platelet aggregation. In platelet-rich plasma, BAEE also inhibited platelet aggregation without inhibiting fibrin clot formation. These results suggest that the marked sensitivity of washed human platelets to small changes in the external pH may lead to misinterpretation of the anti-aggregatory potency of protonated test drugs. For work with human washed platelets a physiological salt solution with more buffering capacity than Krebs-bicarbonate (such as Tyrode-HEPES) is recommended.

Cytomegalovirus infection of gastrointestinal endothelium demonstrated by simultaneous nucleic acid hybridization and immunohistochemistry.

To demonstrate the degree of involvement of endothelial cells in cytomegalovirus (CMV) infection of the gastrointestinal tract we have stained sections from gastrointestinal specimens that showed inclusion bodies on hematoxylin-eosin staining. Factor VIII was first detected using a rabbit anti-factor VIII primary antibody and an alkaline phosphatase-labeled sheep anti-rabbit secondary antibody. The CMV was then visualized with a biotin-labeled CMV probe detected by a streptavidin peroxidase technique with aminoethyl carbazole as the chromogen. Factor VIII staining was a bright blue and CMV a brick red. The specimens included one small-bowel resection and four colonic resections, as well as an esophageal biopsy. The patients' diagnoses included bone marrow transplant recipient, acquired immunodeficiency syndrome, ulcerative colitis, and renal transplant recipient. Cells positive for both CMV and factor VIII ranged from 35% to 60% of positive cells in a representative section, and the relative percentages (mean +/- SE) for cell type for infected cells were: endothelial, 48.9 +/- 4.5; vascular luminal (factor VIII negative), 6.1 +/- 1.7; perivascular (factor VIII negative in vascular wall), 16.2 +/- 3.2; and other cell (non-vascular factor VIII negative), 28.9 +/- 5.1. These findings and clustering of infected cells around the vessels provide evidence that CMV infection of the gastrointestinal tract is primarily vasculitic and related to infection of endothelial cells.

In situ DNA hybridization for cytomegalovirus in colonoscopic biopsies.

We reviewed colonoscopic biopsies of the lower gastrointestinal tract performed during a two-year period. Those representing neoplasia were excluded. Formalin-fixed paraffin-embedded biopsy specimens from 53 patients were studied by in situ DNA hybridization for cytomegalovirus (CMV) using commercially available biotinylated DNA probes detected by an avidin-biotin peroxidase technique. Nine of the patients were severely immunocompromised: four had acquired immunodeficiency syndrome, three had ulcerative colitis and were receiving high-dose steroid therapy, one was a bone marrow transplant recipient, and one had idiopathic pulmonary fibrosis and was receiving therapy with prednisone and cyclophosphamide. Four of these had evidence of CMV infection by routine histology and DNA hybridization. Three additional immunocompromised patients had evidence of CMV infection by DNA hybridization alone. Forty-four patients had inflammatory conditions or ulcerations of the lower gastrointestinal tract. Six of these had evidence of CMV by DNA hybridization alone. Histologically normal as well as enlarged and cytomegalic cells were probe positive, and the cells were sparse to numerous in number. They were found in the epithelium and/or lamina propria. This technique was demonstrated to be applicable to routinely processed colonic biopsy specimens.

A pharmacokinetic and dynamic study of single nightly doses of conventional and controlled release formulations of trazodone.

We studied the pharmacokinetics and dynamics of single evening oral doses of conventional capsules (100 mg) and a controlled release formulation (150 mg) of trazodone in 12 fasting and non-fasting young healthy volunteers. When corrected for the different doses used, there was no significant difference among the areas under the plasma concentration-time curves (AUC) for the conventional capsules and the controlled release tablets under fasting and non-fasting conditions. Both conventional and controlled release formulations were followed by a reduction in critical flicker fusion threshold (CFFT) and this effect was not influenced by the administration of food before dosing. After both conventional and controlled release formulations, blood pressure was significantly lower when medication had been given in the fasting state than when it had been given after food. The frequency of adverse symptoms was greater after the controlled release (150 mg) than after the conventional (100 mg) formulation. We conclude that there is no obvious advantage to the controlled release formulation (150 mg) and that conventional trazodone (100 mg) should be taken after food when it is given at night.

Recent developments in the use of nitrates for treatment of angina pectoris.

Organic nitrates are effective in the treatment and prophylaxis of angina pectoris. The major clinical problem of tolerance may be avoided if the daily plasma concentrations of the active metabolite, isosorbide-5-mononitrate are maintained at 100-300 ng/ml. The most promising development in achieving this is the use of sustained release preparations.