Experimental Biology and Medicine

To chronically decrease activity of the ouabain-sensitive sodium pump in vascular smooth muscle, we administered digoxin (60 μg/kg, followed by 8-16 μg/kg/12 hr, iv) to five male mongrel dogs for 4 weeks. Serum digoxin levels were monitored and maintained at concentrations sufficient to continuously inhibit the sodium pump. Blood pressures and serum Na⁺ and K⁺ concentrations of the treated dogs did not change significantly. Five paired control dogs received saline injections. Two hours after a final injection, samples of small mesenteric arteries and veins were obtained under sodium pentobarbital (30 mg/kg, iv) anesthesia for measurement of vessel wall ⁸⁶Rb uptake and water content. In veins there were no significant changes in pump activity or water content. In contrast, in arteries the ouabain-sensitive ⁸⁶Rb uptake was depressed (P < 0.02) in treated dogs. Accompanying this depression was an increase, averaging 6.2% (P < 0.05), in mesenteric artery wall water content. These data provide evidence associating chronic digitalis suppression of the sodium pump in arteries with the development of wall edema. These results are compatible with the hypothesis that inhibition of the sarcolemmal sodium pump of arteries may underlie the “waterlogging” of these vessels in hypertension.

Busulfan produces profound and prolonged hematopoietic depression in mice. However, the decline in pluripotent cell compartments such as those forming spleen colonies (CFU-S) is much more precipitate than the decline in their progeny such as cells forming colonies in methylcellulose (CFU-C) or in granulocytes. Endotoxin was given immediately following a single 60 mg/kg dose of busulfan in order to determine its influence upon the subsequent decline and recovery of CFU-C, total nucleated cells, and marrow granulocytes/humerus. There was no effect on the rate of decline for 3 days following busulfan. However, the decline slowed in endotoxin treated mice on the 4th day and all three values were significantly higher than in controls on Day 5, the nadir for controls. Controls had an abortive recovery evident by Day 7. At this time endotoxin treated mice had higher values than in controls for total nucleated cells and granulocytes, but not for CFU-C. By Day 9, no further effect of endotoxin could be demonstrated. This effect is similar to that of androgenic steroids on the granulocytic system of busulfan-treated mice. The simplest explanation for the effect is to suggest that CFU-C are capable of self-replication and that endotoxin stimulates this self-replication.

The chemical composition of a “heparin analog” (A73025) was studied and its biologic properties were compared with those of a heparin preparation from intestinal mucosa. A73025 was characterized as a polysulfated chondroitin with a molecular weight of about 10,000 M _r. Unlike heparin, A73025 showed poor anticoagulant activity in vitro by Stypven clotting time, partial thromboplastin time, and thrombin time assays. A73025, like heparin, although to a lesser extent, inhibited thrombin induced platelet aggregation, released lipoprotein lipase when injected into rabbits, and complexed with serum low and very low density lipoproteins.

Hemodynamic, mechanical, and subcellular function in a rat model of chronic ethanol consumption (A) and pair-fed control rats (C) were evaluated over a 17-week course. Between 5 and 17 weeks, isovolumetric peak systolic pressure did not change (C = 260 ± 20 mmHg; A = 265 ± 35 mmHg;p > 0.1). At 17 weeks myocardial contractility as evaluated by (dP/dt-max)/32 x LVP began to decline in the A rats (V_max = 1.46 ± 0.12, r = 0.89; C = 1.560 ± 0.09, r = 0.95) but this is not statistically significant. Neither the velocity of calcium uptake by the isolated sarcoplasmic reticulum (SR) nor the whole heart homogenate (H) was different between C and A rats during the 17-week period and the estimated quantitative content of SR/g Heart did not change (at 17 weeks: mg SR/g Heart = 0.15, C and A). Utilizing Triton X-100 purification of cardiac myofibrils, Mg²⁺-dependent, Ca²⁺-stimulated myofribrillar ATPase activity began to decline at 11 weeks (A = 80% of control) and continued to decline at 17 weeks (A = 72% of control). These data suggest that a depression of contractile protein function may be an early alcohol-induced derangement in myocardial contractility that precedes the onset of significant mechanical dysfunction.

Envelope and core proteins of Venezuelan encephalitis virus were detected by indirect fluorescent antibody staining in cultured primary hamster embryonic cells infected under single-cycle conditions. Antibody to purified core protein stained the entire cytoplasm at 7 hr postinfection, but never the outer surface of the plasma membrane. By 24 hr postinfection, when cytopathic effects were extensive, core protein was present as cytoplasmic granules. Antibody to purified envelope proteins stained a single bright perinuclear area in less brightly fluorescing cytoplasm at 7 hr postinfection, and plasma membranes at 8 hr postinfection. The perinuclear location of Venezuelan encephalitis virus envelope glycoproteins was interpreted as involving the Golgi apparatus which is a known location of intracellular glycoprotein synthesis. Thus, the Golgi apparatus is apparently a source of the smooth membranes with which α-virus envelope proteins have been associated after cell fractionation.

Ferritin-labeled antibodies and lectins have been used to visualize endocytic vesicles in erythrocytes. Distinguishing preformed endocytic vesicles from invaginated plasma membrane has proved difficult especially since antibodies and lectins by themselves induce these changes in membrane configuration. We have used a transmission electron microscope technique (TEM) utilizing positive colloidal iron staining, employing as our model erythrocytes treated with cationic amphipathic drugs. Our technique allows fixation of cells at any point during drug treatment before staining. The membrane lining of unsealed but not of sealed vesicles was coated with electron dense ferric oxide grains. The formation of sealed vesicles is evidence for the fluidity of membranes of the erythrocytes of adults. The data also suggest that the mechanism may not be identical with every cationic amphiphile.

Exhaustive treadmill running caused a significant decrease in hepatic protein levels. The rate of protein loss was 0.23 g/hr. This protein loss was accompanied by an increase in the activities of free (nonsedimentable) cathepsin D and arylsulfatase, suggesting the involvement of lysosomes. A 24-hr fast lowered the liver protein levels by about 20%. Plots of liver protein levels versus the time it took to reach exhaustion showed that the absolute rate of protein loss was the same for both the fed and the fasted rats. Since the initial protein levels were lower in the fasted rats, their relative rate of protein loss was greater. The level of activity of free (nonsedimentable) cathepsin D and arylsulfatase increased as the time to exhaustion increased in both the fed and fasted animals. The increase was severalfold greater in the fasted animals suggesting that fasting alters the structure of lysosomes in a way that enhances the increase in lysosomal fragility caused by exercise.

Adult persons who were given a pneumococcal polysaccharide vaccine containing 50 μg each of 12 serotypes showed an average 10-fold increase in amount of antibody to the 12 antigens (range 6- to 20-fold) 1 month after vaccination and there was an approximate average 50% decline in antibody 31/2 years later. Children who were 2 to 12 years old at the time of vaccination showed about the same antibody response to the vaccine but this was less persistent and there was about a 55% decline, on the average, after only 21 months, The findings are discussed in the light of need for revaccination and of the nature of antibody responses to polysaccharide antigens.

Normal fibroblasts of bearded seal, northern fur seal, sea lion, domestic sheep, and normal kidney cells of spotted dolphin and Pacific common dolphin have been transformed by fragments of SV40 DNA containing the entire early region. The transformed, T-antigen-positive cell strains grew to higher saturation densities and, except for the transformed dolphin kidney cell strains, were able, albeit poorly, to form colonies in soft agar medium. The transformed northern fur seal, spotted dolphin and Pacific common dolphin cell strains have remained diploid, the transformed sheep cells have become hypodiploid due to Robertsonian translocations, and the transformed bearded seal and sea lion cells had become hypotetraploid. Although all the transformed cells have acquired increased divisional potential and prolonged lifespan in cell culture, only the hypotetraploid cell strains of bearded seal and sea lion have evolved into continuous cell strains.

The in vitro reactivity of peripheral blood lymphocytes from burn patients to the T-cell mitogens phytohemagglutinin (PHA) and concanavalin A (Con A) was studied. Sequential studies showed that responses to PHA were either normal or elevated while responses to Con A were below normal and decreased further over the course of time. Analyses of patients' responses revealed no patterns of reactivity of prognostic value. However, the differences in reactivity to the T-cell mitogens PHA and Con A support the hypothesis that thermal injury produces specific rather than general effects on the immune system and lend credence to the concept that burns may exert a preferential influence on functionally distinct subpopulations of lymphocytes.

Bile acid metabolism has been investigated in a newly described animal model depicting juvenile human diabetes (spontaneously diabetic Wistar (BB) rat) and compared to normoglycemic control from the Wistar strain. Diabetic animals used were on insulin treatment except for the last 24 hr. The plasma glucose levels (mg%) of diabetic rat (D) was significantly higher than control rats (C) (150 ± 35 in C vs 340 ± 32 in D). The total bile acid pool (mg/100 g) in D was significantly (P < 0.05) higher when compared to C (9.0 ± 0.8 in C vs 14.9 ± 1.7 in D). The pool of cholic acid was significantly (P < 0.05) increased while that of chenodeoxycholic acid was significantly (P < 0.05) decreased (cholic acid: 5.9 ± 0.45 in C vs 10.06 ± 1.2 in D; chenodeoxycholic 0.90 ± 0.1 in C vs 0.57 ± 0.06 in D). This increased the cholic/chenodeoxycholic acid ratios from 6.6 ± 0.4 in controls to 19.3 ± 2.4 in diabetic rats. These studies have shown diverse alteration in the concentration of the two primary bile acids in the diabetic state.

Samples of abdominal skin from SHR, WKY, and NW rats were examined to determine: (a) if gross structural differences were apparent among the cutanous vascular beds of the three strains of rats, and (b) if the structural differences were generalized or localized to a specific level of the vascular tree. The most significant differences among the three strains of rats were at the level of the smallest (fourth-order) arterioles. The SHR had 30% fewer of the smallest arterioles than WKY and 38% fewer than NW. However, the smallest arterioles from SHR had diameters 22 and 26% larger than similar vessels from WKY and NW, respectively. Consequently, there were no significant differences in calculated resistance among vessels from the three strains of rats. No significant differences were found in the vessel branching angles among the three strains of rats either. It cannot be determined from these studies whether the genetic mechanism responsible for the differences in arteriolar numbers and diameters observed among these highly inbred strains of rats is related to the genetic mechanism responsible for hypertension in SHR.

Blood pressure and renal blood flow were monitored in conscious normotensive and two-kidney one-clip Goldblatt hypertensive dogs. Captopril administered IV in a single dose of 0.1 mg/kg to normotensives increased renal blood flow by 26.8 ± 7.6% and decreased renal vascular resistance, but did not significantly change blood pressure. Cumulative doses of 0.1 and 0.2 mg/kg increased renal blood flow by 29.9± 6.5%, and decreased renal vascular resistance and blood pressure significantly. Qualitatively similar changes in blood flow and vascular resistance of the contralateral kidney were obtained in the hypertensives. Blood pressure was reduced by a mean of 11.8 ± 4.5 mm Hg, renal blood flow increased by 36.2 ± 7.9% by the low dose, blood pressure decreased by 18.4 ± 2.7 mm Hg, and renal blood flow increased by 42.3 ± 11.4% by cumulative doses of the inhibitor. Plasma renin activity was increased by captopril in the normotensives and a greater increase in plasma renin activity was obtained in the hypertensives. The hypotensive and renal vasodilator effects of captopril appear to be related to blockade of the influence of the renin-angiotensin system, but another action, potentiation of kinin-induced vasodilatation, may also be involved.

A specific antibody to human renin was produced employing a pure human renal renin as antigen. This antibody inhibited the activity of human renal renin but did not inhibit the activities of hog renal renin, rat renal renin, and mouse submaxillary gland renin. Using this antibody an immunoaffinity column was constructed. The gel-bound antibody was used to demonstrate its capability to neutralize inactive prorenin in human plasma as well as active plasma renin, indicating that inactive plasma renin contains structures common to renal renin.

Experimental emphysema was induced in eight beagles by repeated administration of crude papain via aerosol and intratracheal instillation. Significant increases in mean linear intercept in the lungs of the papain treated dogs were consistent with morphological evidence of ruptured alveolar septa. The proportions of elastin in the parenchymal connective tissue were calculated from the desmosine and isodesmosine content of the total connective tissue and of purified elastin from the same specimens. Mean pulmonary elastin proportions determined 2 weeks after the last dose of papain in four dogs with experimental emphysema were only 4.52% ± 1.84 whereas mean pulmonary elastin proportions in four control dogs were 14.42% ± 3.55. In two dogs sacrificed 3 months after the termination of the insult, mean pulmonary elastin proportions were not significantly different from those in the control dogs. Since the elastolytic activity of papain is very low these data suggest that the elastin may be degraded in vivo by other mechanisms involving enhancement of endogenous elastase activity.

The effects of psychosocial environmental change upon circulating antibody response to antigenic challenge was investigated in CBA/USC mice. Mice were reared in isolation and selected groups were subsequently exposed to psychosocial stimulation. Antibody titers of mice that remained in isolation were significantly higher than the titers of mice exposed to psychosocial stimulation. One group of mice exposed to psychosocial stimulation and then returned to isolation showed titers significantly below those of mice exposed to psychosocial stimulation only. These data indicate that psychosocial environmental changes can be productive of significant suppression of antibody formation in mice.

Glycerol in hypertonic solution (80 g in 160 ml) was ingested by 36 patients during evaluation for Meniere's disease. One hour after glycerol, serum osmolality rose an average of 5.7% (17 mOsm/liter), and plasma renin concentration fell an average of 41%. Possible mechanisms for suppression of renin release by hypertonic glycerol are discussed.

Cell cultures of simian and human origin infected with two strains each of herpesvirus saimiri (HVS) and herpesvirus ateles (HVA) were compared for production of infectious virus and early and late antigens (EA, LA) in the presence and absence of the tumor promoting agent 12-O-tetradecanoylphorbol-13-acetate (TPA). Second, Epstein-Barr virus (EBV) infected human and simian lymphoblastoid cell lines of high and low cell passages were also compared for enhanced production of early antigen and virus capsid antigen (VCA) using two concentrations of TPA, owl monkey kidney (OMK) and squirrel monkey lung. Cell cultures infected with HVS and HVA with 20 ng/ml of TPA exhibited higher percentages of early and late antigen producing cells and contained 1.0-1.6 logs more virus. Such cells also had earlier cytopathic effects, larger plaques, and a 3-fold increase in the number of plaques. The TPA also enhanced HVS-EA, and LA both in OMK and human skin fibroblast (HSF) cells. The enhancement of EA was approximately 17.0% more in OMK cells and 4.0% more in HSF. The HVS-LA was 22% higher in OMK and 6.0% higher in HSF cells. Pretreatment of OMK cells with TPA prior to HVS or HVA infection showed only a 0.5-0.7 log enhancement of virus. A dose-response of TPA in P₃HRl cells showed that both 20 and 40 ng/ml doses were able to enhance EBV, EA, and VCA significantly with peak enhancement at 40 ng/ml. Higher doses of TPA (60 and 100 ng/ml) resulted in considerable cell death and reduction in antigen production. TPA (20 and 40 ng/ml) stimulated both VCA and EA antigen production in P₃HRl and B95-8 cells, with lesser effects on other human and simian lymphoblastoid cells. However, the stimulation of EA and VCA with these doses of TPA varied for each cell line. Moreover, TPA-treated P₃HRl, B95-8, and 407-I cells on the average also produced 1.0-1.5 logs more virus than the untreated cells. The higher percentage of EA-VCA production from P₃HR1 and B95-8 cells and lower EA-VCA from other human and simian cells suggests that such virus-cell interaction may be influenced by in vitro passages.

Parenteral administration of the potent hepatocarcinogen N-hydroxy-2-acetylaminofluorene to rats results in a substantial inhibition of hepatic RNA polymerase III(C), the enzyme responsible for the synthesis of tRNA and 5 S RNA. The minor component of RNA polymerase III extractable from nuclei shows a 45-65% reduction in activity while the major component of the enzyme extractable from cytoplasm shows a 40-65% inhibition. This reduction of RNA polymerase III activity is sufficient to account for the inhibition of rRNA synthesis produced by administration of the carcinogen in vivo.

The relationship between renal perfusion pressure and renal blood flow (RBF) was studied in the contralateral or not-clipped kidneys of seven two-kidney one-clip renovascular severely hypertensive rabbits and in the right kidneys of eight normotensive rabbits. Thirty to forty days after clipping the left renal artery, severely hypertensive rabbits were characterized by blood pressures significantly greater than normotensive (105 vs 80 mm Hg), and plasma renin activities three times greater than in normotensive rabbits. The absolute RBF to the right kidney was greater in hypertensive rabbits, but these kidneys were hypertrophied so that renal blood flow per gram kidney weight was the same (2.4 ml/min) in kidneys of both hypertensive and normotensive rabbits. Sequentially reducing renal perfusion pressure decreased RBF similarly in both groups suggesting hypertensive rabbits required 27% greater perfusion pressure to achieve RBF similar to that of normotensive rabbits. When the competitive angiotensin antagonist [Sar¹-Ile⁸]-angiotensin II was administered to both groups, there was no significant change in the RBF of normotensives. In the hypertensive rabbits, however, blood pressure dropped to a value not different from normotensive while renal vascular resistance decreased such that RBF was maintained at a normal level. These results suggest the contralateral kidneys of severely hypertensive rabbits chronically adapt RBF to normal levels despite elevated renal perfusion pressure.

The distribution of fluoride and chloride in muscle, liver, and tendon was studied in groups of 6-15 rats sacrificed at 80-240 min after intraperitoneal injections containing radiofluoride (¹⁸F) and radiochloride (³⁶Cl). Liver exhibited a constant ¹⁸F(tissue water):¹⁸F(plasma water) ratio over 120-220 min and this ratio was constant in muscle over 80-120 min. The ¹⁸F_tw/¹⁸F_pw ratios for muscle were lower than those of liver. The ratios of ¹⁸F distribution for both liver and muscle declined significantly at the longest time periods probably as a result of reductions of intracellular pH and/or increases of extracellular pH. Chloride space volumes were determined from chloride analyses of the tissues and plasma and confirmed by the ³⁶Cl(tissue water):³⁶Cl(plasma water) ratios. The fluid space volumes and the ¹⁸F(tissue water):¹⁸F(plasma water) ratios were used to indicate that the average intracellular ionic fluoride concentrations of liver and muscle are, respectively, 79 and 38% that of plasma water. The relative intracellular fluoride concentrations of liver and muscle are related to their respective values of intracellular pH. The entire water of tendon was found to be available to chloride, analytically determined, and to ³⁶Cl and ¹⁸F.

Bacterial L-phase variants may participate in the pathogenesis of some infectious diseases. Previous studies performed in vitro have demonstrated that L-phase variants of Streptococcus faecalis are able to activate both the classical and alternative pathways of complement and that this activation results in death of the organism. The present study was performed in order to determine whether the complement system serves a protective role, in vivo, against L-phase variants of S. faecalis. Mice were depleted of the third component of complement (C3) by treatment with purified cobra venom factor. The C3 depleted mice had a significantly greater magnitude of bacteremia at 1 and 3 hr after the intraperitoneal injection of L-phase variants of S. faecalis than did control animals. Both groups of animals yielded sterile blood cultures at 8 hr. These studies suggest that the complement system participates in the host's defense against L-phase variants of bacteria.

Significant changes in the activities of the enzymes of methionine metabolism occur in the regenerating rat liver. We found that the levels of methionine adenosyl-transferase and cystathionine synthase increased immediately after partial hepatectomy. A secondary peak of activity was noted on the eighth postoperative day. In contrast, activities of both 5-methyltetrahydrofolate homocysteine methyltransferase and betaine-homocysteine methyltransferase declined. This pattern of changes suggests that transsulfuration, rather than methionine conservation, is a major metabolic response in hepatic regeneration.

The effect of a 4-week thiamine deficiency on gluconeogenesis from xylitol, lactate, pyruvate, dihydroxyacetone, and fructose in isolated rat hepatocytes was investigated. Xylitol is metabolized via transketolase, a thiamine-requiring enzyme, whereas the remaining substrates are converted to glucose via enzymes which do not require thiamine. At 7 days, transketolase activity was 81% of the value obtained with hepatocytes isolated from pair-fed control rats and gluconeogenesis from all the above substrates was normal, indicating an excess of transketolase activity present. By 14 days, however, gluconeogenesis was depressed (65-88% of control). At 28 days, both transketolase activity and gluconeogenesis from xylitol were markedly depressed (32 and 73% of control, respectively), but moreover, those substrates not utilizing thiamine-requiring enzymes for gluconeogenesis were equally or more greatly depressed in their gluconeogenic rates (42-70% of control). These data indicate (i) transketolase activity is present in amounts greater than that needed for normal hexose monophosphate shunt operation, and (ii) thiamine deficiency markedly depresses hepatic gluconeogenesis from a variety of substrates, indicating a generalized effect of the absence of this vitamin on metabolism, and more specifically, that the depression in gluconeogenesis from xylitol could be due to a decrease in the activity of non-thiamine-requiring enzymes. The need for caution in interpreting cause-and-effect relationships between enzymes and pathway flux is stressed.

Studies were conducted comparing interferon production of various mouse strains. Methods used included the production of interferon in vivo and in vitro using spleen cell cultures. Strain differences observed in vivo were generally consistent with the results obtained using cell culture methods. In general, high-producing strains in vivo were also high producers using in vitro methods. Strain differences in vivo were greatest using BCG with old Tuberculin (OT) as a challenge. In general, BDF₁ and C57BL/6J mice were high producers while BALB/cByJ and NZB/BLNJ were consistently low responders to both type I and type II interferon inducers. C3H/He and HA/ICR mice could be classified as intermediate interferon producers.

A sensitive enzyme-linked immunosorbent assay (ELISA) for antibodies to double-stranded RNA has been developed using polyinosinic acid . polycytidylic acid (I _n C _n ) as antigen. Bound antibody was detected using alkaline phosphatase-conjugated goat antiimmunoglobulin and ρ-nitrophenylphosphate as substrate. The assay, as demonstrated for rabbit, NZB/NZW mouse, and grivet monkey sera, was far more sensitive and reproducible than quantitative complement fixation. I _n C _n complexed with poly-L-lysine and adsorbed to polystyrene tubes resulted in optimal uptake of antibodies to double-stranded RNA. I _n -C _n without added poly-L-lysine or with other cationic substances adsorbed poorly, as measured by reduced antibody uptake. Specificity of the assay for antibodies to double-stranded RNA was demonstrated by reduction of antibody binding to antigen-coated tubes following adsorption of sera with double-stranded, but not single-stranded RNAs. Similar adsorption studies indicated that this method could be used also to assay for double-stranded RNA. Therefore, a simple ELISA can be used for quantitative determination of either antibodies to double-stranded RNA or double-stranded RNA antigen.

Hepatic lipogenesis was examined during the preweaning and early postweaning period in Zucker fafa and lean rat pups by measuring total in vivo lipogenesis and the activity of two rate-limiting lipogenic enzymes, acetyl-CoA carboxylase (ACC) and fatty acid synthetase (FAS). No significant differences in in vivo lipogenesis, ACC, and FAS activity were present during the preweaning period. After weaning, total lipogenesis and the two lipogenic enzyme activities, ACC and FAS, were significantly elevated in fafa rats compared to lean rats. Thus, early overproduction of lipids by the liver does not appear to be a primary effect of the fa gene.

Low levels of ascorbic acid reversibly inhibit mouse brain and beef heart cyclic nucleotide phosphodiesterase activity in vitro. Inhibition is noncompetitive with a K_t ≃ 10^-4 M. Similar inhibition occurs with D-ascorbate but not with dehydroascorbate or metabisulfite. The kinetics of inhibition by ascorbic acid were similar for “high” and “low” K_m cAMP phosphodiesterase and cGMP phosphodiesterase activity.

The rate of clearance of leukocytic endogenous mediator from the circulation of the rat was determined by measuring the rate of disappearance of activity from the plasma. Leukocytic endogenous mediator was injected iv into groups of rats and the plasma was periodically sampled for the activities that would lower plasma iron, increase plasma fibrinogen, or release bone marrow neutrophils. These activities were assayed by injecting the plasma into groups of normal rats. The t _1/2 's for the clearance of leukocytic endogenous mediator ranged between 6 and 10 min.

The absorptive flux of [2-¹⁴C]urate in the rat proximal tubule was examined using stopped-flow and continuous-flow microperfusion techniques. In the stopped-flow studies, the luminal perfusion was a steady-state equilibrium solution and the luminal contact time was 60 sec. The ratio of disintegrations per minute (dpm) of urate in the reaspirated sample to the initial solution (C/I) was 0.42 ± 0.02 in control experiments. The addition of either phloridzin or ρ-chloromercuribenzoate to the perfusion solution resulted in significantly higher C/I ratios of 0.52 ± 0.03 (P < 0.05) and 0.61 ± 0.006 (P < 0.01), respectively. Substitution of choline for sodium in the perfusion solution or the addition of probenecid did not significantly change the C/I ratio. In continuous-flow microperfusion studies, the fractional absorption of urate from an isotonic saline solution averaged 10.2 ± 0.8%/mm tubule and was significantly inhibited by the addition of probenecid to the perfusion solution. Urate absorption from a steady-state perfusion solution averaged 4.7 ± 1.0%/mm tubule (P < 0.01 compared to the saline solution) and was not affected by the addition of probenecid. These studies indicate that the presence of either phloridzin or ρ-chloromercuribenzoate in the solution bathing the luminal side of the rat proximal tubule cells inhibits urate absorption. Neither the substitution of choline for sodium in the perfusion solution nor the addition of probenecid to the perfusion solution had any significant effect. The failure to demonstrate an effect of probenecid in the stopped-flow studies may have been due to the requirement to use a non-water absorbing solution. When water absorption is permitted to occur, the addition of probenecid to the perfusion solution inhibited urate absorption.

To determine whether the mammalian intestinal tract is capable of reconstituting itself following complete transection, a 1-cm segment of the small or large bowel was excised in 44 rats. In an additional group of 52 rats the colon was transected distal to a diverting colostomy. In each case the open-cut ends of the intestine were simply dropped back into the peritoneal cavity and the abdominal incision was closed with sutures. Eight animals survived simple transection (18%) regaining functional intestinal continuity in 2 weeks and a morphologically normal appearing intestine in 1 month. When the colon was transected distal to a diverting colostomy the survival rate increased almost four-fold with complete reconstitution of the transected segment in 80% of the survivors.

These studies compared calf muscle strength and calf muscle blood flow response to rhythmic exercise, as gauged by postexercise hyperemia, between a group of young adult males (mean age 24.7 years) and an older group (mean age 73.7 years). All subjects were trained to exert maximum voluntary contractions (MVC) of their calf muscles. MVC values averaged 502 ± 16 (SEM) pounds in the young adult group and 362 ± 53 (SEM) pounds in the older group. This difference was significant (P ± 0.05). Calf muscle blood flow was measured by strain gauge plethysmography before and after 3-min bouts of rhythmic isometric calf muscle exercise. Contraction frequency ranged from 20 to 80 contractions per minute (cpm) while the force per contraction ranged from 7.5 to 30% of MVC. In the resting state there were no significant between-group differences in blood pressure or calf muscle blood flow. For each percentage MVC and cpm exercise combination tested, there were no significant between group differences in postexercise hyperemia. These results indicate that the fundamental relationship between MVC and muscle blood flow is not modified as a function of age.

Dose-response curves to angiotensin II were consecutively performed in rats after anesthesia (pentobarbital 60 mg/kg ip), converting enzyme inhibition (SQ14225 1 mg/kg iv), and ganglionic blockade (pentolinium 10 mg/kg sc) to explore the participation of the sympathetic nervous system in the immediate pressor action of the peptide. Experiments were conducted after the animals were maintained on a sodium-deficient diet for 2 weeks. Converting enzyme inhibition was maintained during ganglionic blockade to avoid changes in the circulating level of endogenous angiotensin II which might be caused by the latter maneuver. SQ14225 caused a 35 ± 4 mm Hg (P < 0.001) fall in systolic pressure and a significant shift to the left of the dose-response curve to angiotensin II compared to that obtained after anesthesia alone. In contrast, pentolinium did not further modify responses to angiotensin II despite significantly lowering systolic blood pressure 49 ± 5 mm Hg (P < 0.001). Our data confirm that a major determinant for the response to injected angiotensin II is the prevailing circulating level of the peptide and suggest that a direct effect of angiotensin II on the vascular bed is the most likely mechanism of action in the whole animal, since an intact sympathetic tone is not required for the peptide to evoke its full pressor response.

Previous investigators have reported that the expression of Rh_o(D) antigen activity by human erythrocytes and their membranes depended on the presence of phospholipid. In order to further elucidate the role of phospholipids in expression of Rh_o(D) antigen activity, erythrocyte membranes and partially purified Rh_o(D) antigens were incubated with bee venom phospholipase A_2. Treatment of erythrocyte membranes with phospholipase A₂ resulted in loss of Rh_o(D) antigen activity as detected by hemagglutination inhibition assays. However, subsequent solubilization of these treated membranes with deoxycholate allowed recovery of Rh_o(D) antigen activity. Phospholipase treatment of solubilized Rh_o(D) antigens, which had been partially purified by affinity chromatography on anti-Rh_o(D) IgG agarose columns, did not destroy Rh_o(D) antigen activity. These results suggested that phospholipids did not affect the antigenic determinants of the Rh_o(D) antigen since solubilized, partially purified Rh_o(D) antigens retained their antigenicity following exposure to phospholipase. Phospholipids were presumably required for proper orientation of Rh_o(D) antigens within erythrocyte membranes since Rh_o(D) membranes lost their antigenicity following phospholipase treatment.

The activity of cyclic AMP-dependent protein kinase was studied in heart and aorta of spontaneously hypertensive rats (SHR) and their controls (Kyoto-Wistar). Although the basal activities were comparable, the response to exogenously added, as well as endogenously synthesized, cyclic AMP was significantly decreased in hearts from 5-week-old SHR. This decrease was not observed at the age of 18 weeks. On the other hand, a normal response to exogenous cyclic AMP was observed in the aorta of young animals, while reduced activity was demonstrated in adult SHR. It is conceivable that these biochemical abnormalities are related to disturbances of cardiovascular functions in SHR, thus an increased heart rate is observed at the age of 5 weeks, while normal heart rate and high blood pressure are present in 18-week-old rats. These studies underline the need for careful age related studies of pathophysiological events in spontaneous hypertension in the rat, particularly at the onset of the disease.

The effect of taurine on myocardial contractility was studied utilizing the isolated perfused bullfrog heart. Within the range of 0.5 to 2.8 mM calcium in the perfusion fluid, taurine produced an increase in the developed force of the spontaneously beating isolated perfused bullfrog heart. The effect of taurine was calcium dependent having a large effect at low calcium concentrations but diminishing at high calcium concentrations. Taurine enhanced calcium binding to sarcolemma isolated from bullfrog hearts. The results suggest that taurine produces an inotropic response in the bullfrog heart by increasing calcium binding and calcium influx at the sarcolemma.

Zinc deficiency and essential fatty acid deficiency in rats have gross effects which are very similar. Zinc, prostaglandin E₁, and dihomo-γ-linolenic acid (prostaglandin E₁ precursor) all have very similar effects on rat vascular smooth muscle. Zinc deficiency has no consistent effect on tissue zinc or zinc enzymes. These observations suggest that the mechanism by which zinc deficiency affects growth and metabolism may be via inhibition of prostaglandin synthesis from essential fatty acids. This hypothesis was examined using zinc deficient rats supplemented with oils containing various concentrations of essential fatty acids. Male Wistar rats were maintained on a zinc deficient diet for 5 weeks and supplemented daily with one of three oils containing different concentrations of essential fatty acids: olive oil (mainly oleic acid), safflower oil (mainly linoleic acid), or evening primrose oil (mainly linoleic and γ-linolenic acids). The olive oil treated rats did not benefit from this treatment in any respect. Dermal lesions were actually worse in this group than in the untreated zinc deficient rats. Safflower oil supplementation significantly inhibited the development of dermal lesions but was of only marginal benefit with respect to growth. Evening primrose oil supplementation also blocked the development of dermal lesions and restored growth to 50% of control. It is suggested that a primary defect of zinc deficiency is to inhibit essential fatty acid metabolism to prostaglandins either by blocking linoleic acid desaturation to γ-linolenic acid or by inhibiting mobilization of dihomo-γ-linolenic acid from tissue membrane stores.

In cats receiving continuous intravenous infusion of histamine dihydrochloride (160 μg kg^-1 hr^-1; 40 mg kg^-1 of aspirin was given as a bolus injection either intravenously or intragastrically. When the animals were killed either 1 or 4 hr later, all cats in both groups had ulcers of the antral part of the stomach extending through the mucosa. The mean area occupied by the ulcers was not significantly different in the two groups. Plasma and mucosal salicylate concentrations at the end of the study were not significantly different in the two groups. We conclude that, in cats receiving an intravenous infusion of histamine, intravenous and intragastric aspirin are equally effective in producing antral gastric ulcers.

The course of the disease, HSV-2, inoculated intravaginally in immature mice was ascertained by studying the incidence of vaginitis, encephalitis, and mortality rate. The therapeutic efficacy of various zinc containing medications administered topically 4 hr after infection was studied. It was found that intravaginally applied zinc medicated collagen sponge containing 50 μg zinc sulfate (7H₂O) in acetate buffer, pH 5.5, significantly improved the course of disease and was significantly more effective than zinc administered intravaginally in a cream base. A solution of zinc sulfate administered as a douche was as effective as the comparable concentration of zinc applied through resilient collagen sponge. We conclude that topical administration of zinc through resilient collagen sponge or as a douche used shortly after the infection significantly inhibits the course of the herpes genitalis infection in the mouse.

The effect of metoclopramide, a dopamine antagonist, and L-dopa on plasma aldosterone (PA), plasma renin activity (PRA), and plasma prolactin (PRL) was studied in eight spontaneously hypertensive rats (SHR) and eight Wistar-Kyoto normotensive rats (WKY). Basal PRL levels were greater (P < 0.005) in the SHR (24.3 ± 1.0 ng/ml) than in the WKY (13.7 ± 0.7 ng/ml). Baseline PA in the SHR (49.9 ± 7.2 ng/dl), although higher, were not statistically different from those in the WKY (38.0 ± 7.0 ng/dl). Basal PRA in the SHR (11.0 ± 0.6 ng/ml hr^-1) was not different from that of WKY (11.4 ± 0.8 ng/ml hr^-1). Although both groups of rats displayed significant PA, PRA, and PRL responses to metoclopramide the responses were greater (P < 0.01) in the SHR. These responses to metoclopramide remained exaggerated even after the rats were pretreated with L-dopa. However, administration of L-dopa resulted in similar suppression of all three hormones in the SHR and in the WKY. These findings suggest that there is altered dopaminergic modulation of secretion of PA, PRA, and PRL in the SHR. Alterations in peripheral and central dopamine control of hormone secretion may play a role in the pathogenesis of essential hypertension in the SHR.