bluuebunny/crossref_metadata_2025 huggingface dataset:
Calcium transport in human red blood cells under hypertonic conditions
Authors: Plishker G. A. , Gitelman H. J.
Published: December 2017

Calcium accumulation in intact human erythrocytes is enhanced by incubation in hypertonic solutions. Hypertonicity produces an increased permeability of the membrane to calcium that is time-dependent and occurs in the presence or absence of calcium. When hypertonically treated cells are incubated for more than 30 min in 300 mosmol/kg solutions the permeability of the membrane to calcium returns to basal values. Oligomycin inhibits the effect of hypertonicity on calcium uptake. The inhibitory action of oligomycin diminishes as the external sodium increases and can only be observed when the external concentration of potassium is at or below 3 mM. Low intracellular sodium and high intracellular potassium concentrations increase the uptake of calcium. It is concluded for human erythrocytes that 1) the increased permeability of the membrane to calcium produced by hypertonicity is a time-dependent, reversible phenomenon and is independent of calcium, 2) the increase in intracellular potassium concentration associated with hypertonic exposure is an important factor contributing to this response, and 3) interactions between calcium and components of the sodium-potassium transport system may account for the enhanced uptake of calcium produced by hypertonicity.
Glucose carriers at maternal and fetal sides of the trophoblast in guinea pig placenta
Authors: Yudilevich D. L. , Eaton B. M. , Short A. H. , Leichtweiss H. P.
Published: December 2017

Trophoblast uptake and unidirectional influx of 3H-labeled hexoses were measured relative to L-[14C]glucose (extracellular marker) using a single-circulation, paired-tracer dilution technique. Successive runs were performed in the fetal and maternal circulations of isolated dually perfused guinea pig placentas, obtained from anesthetized dams and perfused for 60--140 min. The leakiness, estimated from the percentage of the L-glucose dose that crossed the trophoblast, varied (25 +/- 3% (SE), n = 28). On the injection side the maximal sugar uptake (Umax) was measured from early venous concentration ratios, since rapid tracer backflux occurred: Umax = (1 -- 3H/14C) x 100. Umax was independent of the leakiness. In all 14 placentas studied, stereospecific saturable transport of D-glucose was demonstrated at fetal (Umax = 56 +/- 4% (SE), n = 14) and maternal (62 +/- 1% (SE), n = 14) surfaces. The mean unidirectional influxes were 3.3 and 3.5 mumol.min-1.g-1, respectively. Uptakes were inhibited by phloretin and less effectively by phlorizin. D-glucose, 3-O-methylglucose, D-mannose and D-galactose had similar Umax values, about four times that of D-fructose. Tracer backflux and transplacental flux were also equal from both sides. It is concluded that similar hexose carriers, which resemble the human erythrocyte carrier, exist at the membrane on both sides of the trophoblast. The nondestructive technique employed characterizes carriers and receptors at the blood side of cells and could be applied to the placenta or other organs in the intact animal.
Calcium action potentials and potassium permeability activation in pancreatic beta-cells
Authors: Ribalet B. , Beigelman P. M.
Published: December 2017

Ionic control mechanisms of mouse pancreatic beta-cell action potentials ("spikes"), in response to glucose, were studied by measuring membrane potentials with intracellular microelectrodes. The curve relating the peaks of the spikes to the log of the external calcium concentration above 10 mM has a slope of 25 mV/10-fold increase of Ca2+. This approaches the value predicted by the Nernst equation for a pure Ca2+ electrode. Increasing the external [Ca2+]o from 0 to 42.5 mM caused an increase in rates of spike depolarization and repolarization. Lowering [Ca2+]o or applying Ca2+ conductance blockers, including Co2+ (1.25 mM), Mn2+ (2mM), and D-600 (2 X 10(-4) M), caused a decrease in rates of spikes depolarization and repolarization, with an increase of [Ca2+]o reversing this effect. Higher concentrations of these Ca2+-conductance blockers eliminated the spike activity. Quinidine at a high concentration (10(-3) M) blocked spike repolarization. Tetraethylammonium (TEA, 25 mM) increased spike amplitude and duration. Therefore, it is concluded that Ca2+ entry during the spike affects potassium permeability, which is inhibited by TEA. Also, there is a competitive binding between Co2+, Mn2+, Mg2+, and Ca2+, the charge carrier. These cations may have an additional action of substituting for Ca2+ to "stabilize" the membrane.
Characteristics of calcium transport and binding by rat myometrium plasma membrane subfractions
Authors: Grover A. K. , Kwan C. Y. , Crankshaw J. , Crankshaw D. J. , Garfield R. E. , Daniel E. E.
Published: December 2017

A gradient has been designed to yield two subfractions of plasma membrane vesicles from rat myometrium, a low buoyant density (8-24% sucrose) fraction N1 richer in 5'-nucleotidase and a higher buoyant density (24-30% sucrose) fraction N2, instead of a previously described fraction F1. Both N1 and N2 had very low activities of NADPH-cytochrome c reductase and succinate-cytochrome c reductase. Electron micrographs of thin sections of N1 showed clear vesicles, whereas N2 consisted of vesicles with electron-dense bodies attached to them. These plasma membrane vesicles can actively take up Ca. The active uptake of Ca was potentiated by oxalate and phosphate and abolished by the Ca ionophore A23187. Dilution of actively loaded vesicles in isotonic media containing EGTA led to loss of a small proportion of the stored Ca instantaneously and the remainder more slowly in a biphasic manner. Dilution in hypotonic media with EGTA led to a release of a much larger proportion of the accumulated Ca. A23187 at high concentrations (10 microM) caused a release of all the sequestered Ca whether the active Ca uptake had been carried out in the presence or in the absence of oxalate. A23187, 0.5 microM, released all the sequestered Ca from the vesicles that were actively loaded in the absence of oxalate, but only 37% when the vesicles were actively loaded with Ca in the presence of oxalate. Comparison of the composite plasma membrane fraction F1 (8-30% sucrose) and the subfractions N1 and N2 showed that they had different capacities for Ca uptake in the presence and absence of ATP. An attempt has been made to analyze the active Ca-uptake data in terms of various Ca pools.
Gap junction formation and regulation in myometrium
Authors: Garfield R. E. , Merrett D. , Grover A. K.
Published: December 2017

Myometrial tissues from pregnant rats were examined by electron microscopy for the presence of gap junctions after incubation in vitro with a variety of agents. Gap junctions were present in low frequency or absent prior to incubation in vitro. The junctions were present in control tissues in high frequency after 48 h incubation. The addition of cycloheximide or actinomycin D inhibited the incorporation of [3H]leucine into TCA-precipitable proteins and prevented gap junction formation. A prostacyclin analog (carbacyclin), a thromboxane synthesis inhibitor, and indomethacin also prevented gap junction formation. Oxytocin had no effect on gap junction formation but isoxsuprine decreased their number and increased their size. Isoxsuprine and isoproterenol also produced electron opaque crystals associated with the gap junctions. Dibutyryl cAMP treatment but not monobutyryl cGMP also increased the size of gap junctions. Based upon this and previous studies, we propose at least four sites for regulation of gap junctions and possible control of labor.
Dependence of membrane potential on extracellular ionic concentrations in myotonic goats and rats
Authors: DeCoursey T. E. , Bryant S. H. , Owenburg K. M.
Published: December 2017

Intracellular potassium concentration([K]i) and the resting sodium-to-potassium permeability ratio (alpha = PNa/PK) were determined by recording resting potentials at 25 degrees C in six solutions with variable potassium concentrations, constant [K+] + [Na+], and constant [K+] X [Cl-] product and fitting the results by nonlinear least squares to the Goldman-Hodgkin-Katz equation. Fits to models including chloride permeability terms suggest that chloride is in Donnan equilibrium in the muscles studied. External intercostal muscle was biopsied from anesthetized normal goats and goats with hereditary myotonia. In normal goat muscle, alpha was 0.012 +/- 0.001 (mean +/- SE, n = 11), and [K]i was 133 +/- 6 mM (n = 9); myotonic goat muscle did not differ from this. Extensor digitorum longus (EDL) muscle was removed from anesthetized male Wistar rats and from like rats pretreated with a single large dose of 20,25-diazacholesterol (DAC). In normal rat EDL, alpha was 0.019 +/- 0.002 (n = 12), and [K]i was 142 +/- 4 mM (n = 12). In the 16-day DAC-treated rats, alpha was significantly reduced (P < 0.002) to 0.010 +/- 0.002 (n = 8), and [K]i was slightly reduced by DAC. It is concluded that DAC reduced PNa in rat fibers. Tetrodotoxin at 5 microM did not affect either parameter in the rats.
Effects of lactic acid accumulation and ATP decrease on muscle tension and relaxation
Authors: Sahlin K. , Edstrom L. , Sjoholm H. , Hultman E.
Published: December 2017

The present study was undertaken to evaluate the effect of lactic acid accumulation on peak tension and relaxation rate of the isometric twitch. Isolated extensor digitorum longus muscle from rat was stimulated electrically at a frequency of 2/s under anaerobic conditions. Comparison was made with muscles in which glycolysis was blocked with iodoacetic acid (IAA). Stimulation of unpoisoned muscles for 3 min decreased tension to 50% and increased relaxation time to 250% of the initial value. Lactate increased 15-fold, and muscle pH decreased from 7.10 to 6.76. Stimulation of IAA-poisoned muscles for 1 min decreased tension to 50% but did not increase the relaxation time. Stimulation of IAA-poisoned muscle resulted in a pronounced decrease (about 50%) of the ATP and total adenine nucleotide content in muscle, whereas only a small decrease (10-15%) occurred in unpoisoned muscle. The main findings in the present study were that tension decline in unpoisoned muscle is closely related to decrease in muscle pH and increase in ADP but not to ATP content per se and that slowing of relaxation rate is closely related to decrease in muscle pH but not to muscle content of ATP or creatine phosphate.
Formation of stimulus-induced vacuoles in serous cells of tracheal submucosal glands
Authors: Mills J. W. , Quinton P. M.
Published: December 2017

Parenchymal cells of exocrine glands have been reported to develop numerous large vacuoles when intensely stimulated either pharmacologically, electrically, or physiologically. We have found that tracheal submucosal glands of the cat form similar vacuoles in response to alpha-adrenergic stimulation. Light and electron microscope examination of tracheal submucosal glands stimulated in vitro with methoxamine (10(-4) M) reveals that only serous cells of the gland acini develop vacuoles. Ductal cells and mucous acinar cells do not form vacuoles. Furthermore, several lines of evidence indicate that within serous cells vacuoles arise from secretory granules, which apparently swell as they accumulate fluid during intense stimulation. The following paper describes some of the factors and characteristics of stimulus-induced vacuolation.
Deleterious effects of calcium deprivation on freshly isolated hepatocytes
Authors: Edmondson J. W. , Bang N. U.
Published: December 2017

Biochemical and morphological properties of rat hepatic parenchymal cells isolated without calcium were compared to cells isolated by adding calcium to the isolation medium at the time of addition of collagenase. Calcium contents of the two cell preparations were 4.5 +/- 0.3 and 10.5 +/- 0.5 nmol/mg dry wt, respectively (P les than 0.001). Magnesium content of both preparations was 37 nmol/mg dry wt. Potassium contents were 92 and 154 meq/l, respectively (P less than 0.001). Potassium content of calcium-deficient cells increased to 161 meq/l following incubation for 30 min in a medium containing 1.6 mM ionized calcium. When incubated in a medium containing a subphysiologic concentration of ionized calcium, calcium-deficient cells rapidly lost the ability to exclude trypan blue and to retain lactate dehydrogenase activity. As contrasted to calcium-sufficient hepatocytes, calcium-deficient cells failed to accumulate alpha-aminoisobutyric acid by active transport and lacked microvilli and nuclear contents. This study supports simultaneous addition of calcium and collagenase to the isolation medium as a means for preserving physical, functional, and morphological integrity of isolated hepatic parenchymal cells.
Uptake and asymmetric efflux of amino acids at maternal and fetal sides of placenta
Authors: Eaton B. M. , Yudilevich D. L.
Published: December 2017

Unidirectional uptake of eighteen amino acids into the syncytiotrophoblast was measured from both the maternal and fetal circulations of isolated dually perfused guinea pig placentas using a single-circulation, paired-tracer dilution technique. A bolus containing a tritiated amino acid and L-[14C]glucose (extracellular marker) was injected intra-arterially into one circulation, and both venous outflows were sequentially sampled. The maximal cellular uptake (Umax) on the injection side was determined from (1-[3H]/[14C]) values and used to calculate the unidirectional influx. Umax values for neutral and basic amino acids ranged between 15 and 58% and were similar on both sides of the trophoblast. Uptake of the acidic amino acids and taurine was minimal. Amino acid influx from either circulation was followed by rapid tracer backflux and transplacental transfer. Tracer efflux was asymmetric and preferentially directed towards the fetal side. It is suggested that amino acid transport systems are present on both surfaces of the placenta and that net transfer from mother to fetus is the result of asymmetric efflux from the trophoblast.
Mechanical properties of tenotomized and denervated-tenotomized muscles
Authors: Dasse K. A. , Chase D. , Burke D. , Ullrick W. C.
Published: December 2017

Mechanical properties of rat soleus and plantaris muscles were studied in vitro following tenotomy, denervation, or tenotomy plus denervation, all of 3 wk duration. Controls included muscles from sham-operated animals and from animals with muscle tendons severed but immediately resutured. Results of twitch times, times to peak tension, and times to half-relaxation for 145 muscles clearly showed that the slight increase in muscle speed that occurs in the soleus muscles only is due to severance of the muscle tendon per se and not related to muscle shortening and possible related alterations in muscle spindle activity that occurs in simple tenotomy. Furthermore, any demonstrable mechanical changes that occur with tenotomy or with section and resuturing of the tendon requires the presence of intact innervation. We conclude that, contrary to published opinion, tenotomy does not transform slow contracting muscles into fast contracting muscles and that comments concerning the role of stretch receptors in this postulated transformation are pure conjecture, unsupported by experimental data.
Hyperpolarization of mammalian skeletal muscle fibers in K-free media
Authors: Akaike N.
Published: December 2017

Time course of changes in resting potential following removal of K ions from Krebs solution was studied in the isolated rat soleus muscle. In K+-free Krebs solution the muscle fibers hyperpolarized to a peak within 60 min followed by a variable, gradual depolarization over the next few hours. A significant decrease of the intracellular K+ concentration ([K]i) occurred within 30 min in K+-free Krebs. Compensatory increase of the intracellular Na+ concentration ([Na]i) lagged behind [K]i change. Both hyperpolarization and successive depolarization were modified in K+-free medium containing different [Na]o, [Ca]o, or osmotic strength. A part of the hyperpolarization was reduced by adding ouabain. Therefore, ouabain-sensitive potential of the fibers in K+-free medium was attributed to the activation of electrogenic Na pump by K+ leaked out continuously from the fibers and accumulated just outside of cell membranes but not to changes of membrane Na+ and Cl- permeabilities. It is concluded that hyperpolarization of muscles exposed to K+-free medium is the sum of the diffusional K+ potential and ouabain-sensitive metabolic potential. Interestingly, the time course of changes of membrane response to K+-free krebs was identical for contralateral soleus muscles in the same rat, though it varied among different rats.
Development of contractile properties in avian embryonic skeletal muscle
Authors: Reiser P. J. , Stokes B. T.
Published: December 2017

The development of the twitch and tetanic responses of the embryonic chick posterior latissimus dorsi muscle has been studied during the last week in ovo. Normalized twitch and tetanic forces increased 3- and 12-fold, respectively, during this period. The changes in the kinetics of the twitch and tetanic responses differed during this developmental period. The time to peak twitch force progressively decreased. The decrease in time to half-peak tetanic force and the increase in the time differential of force production of the tetanic response did not continue after day 18. A prolonged tonic contractile component was described for both the twitch and tetanic responses, particularly in muscles from the younger embryos (days 14-18). A large decrease in the time to one-half relaxation of the twitch response also takes place during the final week in ovo. This detailed description of the development of the contractile properties provides a model system of fast-twitch muscle development in which neurogenic and myogenic components of muscular differentiation can be studied from several approaches.
Amiloride: a molecular probe of sodium transport in tissues and cells
Authors: Benos D. J.
Published: December 2017

The potassium-sparing diuretic amiloride has proven to be a useful pharmacological tool for elucidating the molecular basis and physiological regulation of facilitated sodium entry in tissues and cells. There are two general classes of Na+ transport mechanisms which are sensitive to this drug: 1) a conductive Na+ entry pathway found in electrically high resistance epithelia and 2) a Na+-H+ electroneutral exchange system found in certain leaky epithelia such as the renal proximal tubule. This latter system is also found in many different cellular preparations and seems to function in cell proliferation and differentiation, volume regulation, and intracellular pH regulation. In these cells, this exchange pathway becomes operational usually after some external stimuli. Much higher concentrations of amiloride are required to inhibit the exchange pathway than those required to inhibit the Na+ entry pathway. This drug is the most potent and specific inhibitor of Na+ entry found to date and thus affords the opportunity to be used as a label for the isolation of these transport moieties.
Role of the motor nerve in activity-induced enzymatic adaptation in skeletal muscle
Authors: Henriksson J. , Galbo H. , Blomstrand E.
Published: December 2017

The sciatic nerve was cut on one side in 11 male cats, and a piece of the nerve was removed. The cats were then divided at random into two groups, a stimulation group (S) of five cats and a control group (C) of six cats. Bilateral electrical stimulation (2 Hz) of the gastrocnemius muscle (directly or via the motor nerve) was carried out in the S cats 4 h/day, 3 days/wk for 4 wk. The voltage delivered was adjusted in each cat so that both gastrocnemius muscles lifted identical loads the same distance. The activity of the tricarboxylic acid cycle marker enzyme succinate dehydrogenase (SDH) per unit of muscle weight more than doubled in response to stimulation both in the intact and the denervated gastrocnemius muscle. Stimulation did not affect the activity of the glycolytic marker enzyme 6-phosphofructokinase (PFK) or muscle capillarization. Denervation resulted in pronounced (approx 50%) fiber atrophy, which was not prevented by the stimulation. It is concluded that the presence of the motor nerve per se is not necessary for an activity-induced adaptation of the oxidative capacity of skeletal muscle.
Effects of 1,25-(OH)2D3 administered in vivo on phosphate uptake by isolated chick renal cells
Authors: Liang C. T. , Barnes J. , Cheng L. , Balakir R. , Sacktor B.
Published: December 2017

Renal cells from Vitamin D-deficient and 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3]-repleted chicks were isolated by a collagenase-hyaluronidase procedure. Exclusion of trypan blue and respiratory measurements indicate that the cells were functionally intact and metabolically active. The uptakes of phosphate and alpha-methylglucoside were stimulated markedly by Na+ in the extracellular medium. Phosphate uptake in the presence of Na+ was saturable with respect to phosphate concentration; half-maximal activity was obtained with approximately 0.2 mM. Three hours after 1,25-(OH)2D3 was injected into vitamin D-deficient chicks the Na+-dependent phosphate uptake by the isolated cells had increased about 40%, i.e., 2.00 compared with 1.44 nmol.min-1.mg protein-1. Phosphate uptake in the presence of K+ in the extracellular medium and alpha-methylglucoside uptake in the presence or absence of Na+ were unchanged. In a secondary response found 17 h after 1,25-(OH)2D3 injection, Na+-dependent phosphate uptake decreased. Serum concentrations of phosphorus and calcium were not measurably changed in the 3-h repleted bird, but both levels were increased 17 h after treatment. Administration of phosphate into vitamin D-deficient chicks, so that the serum concentration of phosphorus was raised to that of the 17-h 1,25-(OH)2D3 repleted animal, effected a comparable decrease in phosphate uptake. Serum calcium levels were not altered by this treatment. The actions of parathyroid hormone in stimulating adenylate cyclase and in inhibiting phosphate uptake were notably blunted in the vitamin D-deficient chick. Sensitivity to parathyroid hormone was not restored until several days after 1,25-(OH)2D3 repletion. These findings suggest that the initial response to 1,25-(OH)2D3, to increase renal phosphate uptake, and the secondary response, to decrease phosphate uptake, were by parathyroid hormone-independent processes. The results also indicate that the isolated renal cell represents an excellent model for studying the mechanism by which 1,25-(OH)2D3 regulates phosphate transport in the kidney.
Type IIB to IIA fiber transformation in intermittently stimulated rabbit muscles
Authors: Mabuchi K. , Szvetko D. , Pinter K. , Sreter F. A.
Published: December 2017

Long-term intermittent stimulation (10 Hz, 8 h/day, 7 wk) of the fast-twitch tibialis anterior results in a complete transformation of type IIB fibers to type IIA fibers. This is shown by the histochemical ATPase reaction and by a decrease in Ca2+-uptake ability by the sarcoplasmic reticulum. Furthermore, as shown by studies on bulk myosin and on single fibers, the LC1-to-LC3 light chain ratio is increased on sodium dodecylsulfate gel electrophoretograms, and there are changes in the myosin isozyme pattern manifested on pyrophosphate gels under nondissociating conditions. Thus the staining intensity of the slower moving putative LC1 homodimer band increases, and there is a difference in migration velocity between stimulated and unstimulated isozymes suggesting a possible difference in the heavy chain. This study underlines the importance of the stimulation schedule in determining whether a fast-to-slow transformation or a shift in subtype takes place.
Structure of tight junctions during Cl secretion in the perfused rectal gland of the dogfish shark
Authors: Forrest J. N. , Boyer J. L. , Ardito T. A. , Murdaugh H. V. , Wade J. B.
Published: December 2017

In epithelia that secrete sodium chloride, high-conductance tight junctions between cells have been proposed as the primary pathway for transepithelial sodium flux. We examined the properties of tight junctions in the perfused rectal gland of the dogfish shark during basal secretion and following adenosine 3',5'-cyclic monophosphate stimulation of sodium chloride secretion. Freeze-fracture electron microscopy revealed extensive interdigitation of adjacent cells with an associated amplification in the length of tight junctions per area of luminal surface, averaging 102 +/- 4.7 m/cm2 in outer regions of 80 +/- 6.7 in inner regions of the gland. Marked heterogeneity of junctional structure was present with junctional elements varying from single strands to three duplex elements and junctional depth varying from 15 to 60 nm. In glands perfused with lanthanum chloride, ionic lanthanum filled the intercellular space up to but not through the tight junctions. Characteristics of tight junctions were not different during basal and maximally stimulated sodium chloride secretion. These studies define tight junctions in the rectal gland as an anatomical barrier capable of restricting the passage of relatively small molecules such as urea while providing a greatly amplified junctional area for the passive diffusion of sodium and water.
Production of ornithine by intact human erythrocytes
Authors: Williams J. A. , Phang J. M.
Published: December 2017

Although the conversion of arginine to ornithine in humans is largely an hepatic process, erythrocytes also have this enzymatic capacity. We have shown that intact human erythrocytes readily produce ornithine from arginine and release product ornithine to the incubation medium. The rate of ornithine formation from arginine was 0.22 mumol.h-1.ml cells-1, and 85% of the ornithine is recovered extracellularly. Moreover, we have shown that ornithine and urea are the unique and stoichiometric products of arginine metabolism in the intact erythrocyte. The rate of ornithine production by intact red blood cells was a saturable function of arginine concentration in the medium; the derived Km for arginine for this conversion was 0.16 mM, a value striking for its close approximation to physiological arginine concentration in human plasma. We propose that the production and delivery of ornithine by intact red blood cells may supply peripheral tissues such as bone and muscle with an important precursor for proline and polyamines.
Insulin stimulation of protein synthesis in cultured skeletal and cardiac muscle cells
Authors: Airhart J. , Arnold J. A. , Stirewalt W. S. , Low R. B.
Published: December 2017

The effects of acute exposure to insulin on protein synthesis were examined in primary, differentiated cultures of embryonic chick heart and skeletal muscle cells. Synthetic rates were calculated using the specific activity of tRNA-bound leucine as precursor, a specific activity that was significantly less than that of extracellular leucine but greater than that of free, intracellular leucine at 0.2 mM external leucine. Insulin did not alter these relationships. Doses of insulin in the physiological range produced significant stimulation of protein synthesis in both cell types. Maximal responses, involving approximately 30% increases in both absolute and fractional rates, were observed at higher insulin concentrations. Significant stimulation by insulin was seen in cardiac cells after only 1 h of insulin treatment, and the effects of the hormone were observed both in the presence and absence of serum in the culture medium.
Sympathetic supraspinal control of venous membrane potential in spontaneous hypertension in vivo
Authors: Willems W. J. , Harder D. R. , Contney S. J. , McCubbin J. W. , Stekiel W. J.
Published: December 2017

To clarify the mechanisms controlling mesenteric venous membrane potential (Em) in spontaneously hypertensive rats (SHR) and Wistar-Kyoto normotensive rats (WKY), Em was measured in vivo using flexibly mounted glass microelectrodes. Venous diameters were also measured. Ems under control conditions were significantly less in SHR. alpha-Adrenergic blockade with phenoxybenzamine (PBZ) significantly dilated and hyperpolarized SHR small veins but failed to alter the membrane potential of WKY small veins. After blockade with PBZ, membrane potentials were similar in both strains. Propranolol suffusion failed to alter the SHR membrane potential but depolarized WKY small veins. During blockade with PBZ, propranolol depolarized venous membranes equally in both strains. Membrane potentials after spinal section were similar in both strains and equal to the Ems during total adrenergic blockade. The data indicate that, in SHR, the relatively depolarized Em derives from altered alpha-adrenergic input. In SHR, alpha-adrenergic input appears to exert its depolarizing effect, at least in party, by interfering with the expression of endogenous, beta-adrenergic hyperpolarization. The beta-adrenergic influence, like the alpha-adrenergic input in SHR, is dependent on intact supraspinal pathways.
Cost of force development as a function of stimulus rate in rat soleus muscle
Authors: Loiselle D. S. , Walmsley B.
Published: December 2017

The energy cost (determined myothermically) and mean stress (force per cross-sectional area) development of soleus muscles from adult rats were determined. Muscles were stimulated at various frequencies from 0.25 to 100 Hz over a 4-s interval, and the resulting stress-time integral was averaged over this period to yield a total heat vs. mean stress relation. This relation was characterized by three zones, i.e., 1) a linear zone corresponding to unfused twitches, 2) an intermediate zone corresponding to partial fusion during the relaxation phase only, and 3) a second linear zone corresponding to partially and fully fused tetani. The energy cost of mean stress development in the twitch zone was about four times higher than that of the fusion zone. The results are discussed in terms of the firing frequencies at which motor unit recruitment occurs in mammalian slow-twitch muscles.
Effect of glutaraldehyde on hydrosmotic response of toad bladder to vasopressin
Authors: Eggena P.
Published: December 2017

The present study investigates the time-, dose-, and temperature-dependence of glutaraldehyde action on the permeability to water of the toad bladder. Bladders preincubated with increasing concentrations of glutaraldehyde become progressively desensitized to the hydrosmotic action of vasopressin (ADH), theophylline, and dibutyryl adenosine 3',5'-cyclic monophosphate (dibutyryl cAMP). The ADH response was reduced by 50% with 0.03% glutaraldehyde applied to the serosal side for 10 min at 4 degrees C. Sixfold higher doses of glutaraldehyde were required with mucosal application. Bladders partially fixed with low-dose glutaraldehyde exhibit a markedly prolonged duration of action of ADH. Bladders fixed with higher doses of glutaraldehyde in the presence of ADH retain a high permeability to water for prolonged periods even in the absence of ADH. This action of glutaraldehyde to stabilize the hormone-induced water channels is also considerably more effective with serosal than with mucosal application. As the rate-limiting permeability barrier for water affected by ADH is known to be located in the apical membrane, these findings suggest that glutaraldehyde exerts its action from an intracellular position. It is postulated that glutaraldehyde stabilizes the ADH-induced channels by cross-linkage of amino groups and other reactive sites at the cytoplasmic surface of the apical membrane and/or by inactivating the intracellular machinery responsible for the dispersal or removal of water channels in the hormone target cell.
Differential response to hormones of defined distal nephron epithelia in culture
Authors: Wilson P. D. , Horster M. F.
Published: December 2017

Defined cultures of rabbit kidney cortical collecting tubule (CCT) and cortical thick ascending limb of Henle's loop (CAL) were grown in monolayers from individual microdissected tubules and maintained for up to five passages, a maximum of 53 days. CCT cells contained cytochemically demonstrable vasopressin-stimulated adenylate cyclase, whereas CAL cells were characterized by the localization of Na+-K+-ATPase. [3H]thymidine labeling index decreased with time in primary cultures in the presence or absence of 3% serum. When added to unsupplemented serum-free media alone or in combinations, the growth factors dexamethasone, thyroxine, insulin, epidermal growth factor, and prolactin stimulated [3H]thymidine incorporation to different extents. CCT cells were maximally stimulated by addition of dexamethasone alone, whereas a combination of dexamethasone, thyroxine, insulin, and prolactin was most stimulatory for CAL cells. Addition of hormones concerned with renal ion and water transport to fully supplemented serum-free media inhibited [3H]thymidine labeling index: 1) vasopressin, isoproterenol, and dibutyryl cAMP were equally inhibitory in CCT and CAL cultures; 2) parathyroid hormone and prostaglandin E1 were more inhibitory in CAL cultures; and 3) aldosterone was particularly inhibitory in CCT cultures.
Pancreatic beta-cell electrical activity: the role of anions and the control of pH
Authors: Eddlestone G. T. , Beigelman P. M.
Published: December 2017

The influence of chloride on the mouse pancreatic beta-cell membrane potential and the cell membrane mechanisms controlling intracellular pH (pHi) have been investigated using glass microelectrodes to monitor the membrane potential. It has been shown that chloride is distributed passively across the beta-cell membrane such that chloride potential is equal to the membrane potential. Withdrawal of perifusate chloride or bicarbonate and the application of the drugs 4-acetamido-4'-isethiocyanostilbene-2,2'-disulfonic acid (SITS) and probenecid, both blockers of transmembrane anion movement, have been used to establish that a chloride-bicarbonate exchange system is operative in the cell membrane and that it is one of the control mechanisms of pHi. Amiloride, a specific blocker of the transmembrane sodium proton exchange, has been used to demonstrate that this mechanism is also operative in the beta-cell membrane in the control of pHi. The hypothesis that the calcium-activated potassium permeability is proton sensitive at an intracellular site, a fall in pHi causing a fall in permeability and an increase in pHi causing an increase in permeability, has been used to explain many of the effects observed in this study.
Calcium flow-independent actions of calcium channel blockers in toad urinary bladder
Authors: Levine S. D. , Levin D. N. , Schlondorff D.
Published: December 2017

A role for transmembrane calcium movement in vasopressin stimulation of its target cell has been postulated based on studies with calcium entry blockers such as verapamil. We examined the effect of three sets of structurally different calcium blockers--D600 (an analogue of verapamil), diltiazem, and nifedipine--on water flow in toad bladder. D600 (200 microM), diltiazem (200 microM), and nifedipine (60 microM) inhibited vasopressin-induced water flow but enhanced adenosine 3',5'-cyclic monophosphate (cAMP)-induced water flow, suggesting that the drugs inhibit cAMP generation in response to vasopressin but enhance the response to exogenous cAMP by inhibiting phosphodiesterase activity. In the case of vasopressin stimulation, inhibition of cAMP generation appears to be the overriding effect. This was confirmed by measurements of cAMP content and the protein kinase ratio (-cAMP/+cAMP), which were significantly lower in bladders receiving both D600 and vasopressin than in those receiving vasopressin alone. Furthermore the drugs inhibited activation of adenylate cyclase by vasopressin in cell homogenates and inhibited phosphodiesterase in both homogenates and membrane-free supernatants. Thus these "calcium channel blockers" can directly alter cAMP metabolism in settings where movement of calcium should be irrelevant. The close correlation between the biochemical and transport effects of these agents suggests that their effect on water flow may occur by a direct effect on cellular enzymes or the membranes in which they reside and not by altering local calcium concentrations.
Passive calcium movements in dog red blood cells: anion effects
Authors: Parker J. C.
Published: December 2017

Calcium influx in dog red blood cells was stimulated by replacing chloride in the medium with nitrate or thiocyanate. These anion effects were due to stimulation of a sodium-dependent calcium pathway, because calcium influx in the presence of nitrate or thiocyanate was 1) inhibited by external sodium, 2) dependent on internal sodium, 3) inhibited by cell shrinkage and low pH, and 4) inhibited by quinidine. All these characteristics had previously been shown to hold for calcium movements in the presence of chloride. Neither nitrate nor thiocyanate caused an irreversible change in calcium transport in the concentrations studied. Calcium-stimulated sodium efflux is stimulated when chloride is replaced by thiocyanate but not by nitrate. Several limiting features of the system are discussed, which preclude a conclusive interpretation of the data. The possibility is considered that the rates of sodium-dependent calcium transport in the presence of chloride, nitrate, and thiocyanate are a function of the conductance of these anions.
K+-stimulated sugar uptake in skeletal muscle: role of cytoplasmic Ca2+
Authors: Valant P. , Erlij D.
Published: December 2017

We have compared the effects of insulin with those of elevated external K+ concentration ([K+]o) on sugar uptake and protein synthesis by frog skeletal muscle. When [K+]o was between 0.5 and 15 mM there were no effects on uptake; however, when 20 mM was used a significant increase was observed. Further increases in [K+]o caused larger stimulations of uptake. The stimulation persisted for 2.5 h after washing the high [K+]o. The stimulations caused by both insulin and high K+ were markedly inhibited by cytochalasin B. Dantrolene nearly abolished the response to high K+, whereas it had only minor effects on the resting sugar uptake and on the stimulations caused by either insulin or epinephrine. These results suggest that while both insulin and high K+ activate the cytochalasin-sensitive transport system of sugar transport, each agent must act through a different pathway, because only the effects of high K+ were dantrolene sensitive. The effect of dantrolene suggests that the enhancement of sugar transport caused by high K+ is due to an increase of cytoplasmic Ca2+. In contrast to insulin, high K+ did not modify the rate of protein synthesis.
Ca2+ regulation of tight-junction permeability and structure in Necturus gallbladder
Authors: Palant C. E. , Duffey M. E. , Mookerjee B. K. , Ho S. , Bentzel C. J.
Published: December 2017

To explore the role of Ca2+ in tight-junction permeability, the Necturus gallbladder was exposed to varying Ca2+ concentrations and to the Ca2+ ionophore A23187 added to the mucosal side (1.9 X 10(-6) to 6.8 X 10(-5) M). Electrophysiological parameters measured in an Ussing-type chamber were correlated with tight-junction morphology revealed by freeze-fracture electron microscopy. In Ca2+-free bathing media, transepithelial resistance decreases and tight-junctional ultrastructure is fragmented. In 1.8 mM Ca2+ media, A23187 induces an initial drop in transepithelial resistance, followed by an increase in transepithelial resistance to a value 20% above base line. At peak response to A23187, NaCl diffusion potentials decrease. Freeze-fracture replicas reveal that the number of junctional strands increase pari passu with junctional depth. Both physiological and morphological changes were partially reversible. The initial decrease in transepithelial resistance coincided with a persistent hyperpolarization of the mucosal cell membrane potential difference and a decrease in the mucosal-to-serosal cell membrane resistance ratio. Thus A23187 alters both the transcellular and paracellular pathway, resulting in opposing effects on transepithelial resistance.
beta-Adrenergic blockade of prostaglandin E2- and D2-induced erythroid colony formation
Authors: Belegu M. , Beckman B. , Fisher J. W.
Published: December 2017

beta-Adrenergic receptors have been linked to the actions of beta-adrenergic agonists as well as that of other hormones on erythroid cells. In the present studies, arachidonic acid, the precursor for the endoperoxide intermediates for prostaglandins, was demonstrated to produce a significant increase in erythroid colony (CFU-E) formation in normal mouse bone marrow cultures. Meclofenamate, a cyclooxygenase inhibitor drug that inhibits prostaglandins synthesis, significantly inhibited the increase in CFU-E colony-forming cells produced by arachidonic acid, thus establishing that arachidonic acid was probably converted to some prostaglandin or prostaglandin metabolite in the bone marrow to trigger CFU-E. Prostaglandins E2 (PGE2) and D2 (PGD2), both of which have been demonstrated to be produced in the bone marrow, were found in the present studies to increase the number of CFU-E colonies in normal mouse bone marrow cultures. DL-Propranolol, a beta 1, beta 2-adrenergic blocking agent, and D-propranolol, a non-beta-blocking isomer with nonspecific membrane stabilizing effects, both produced a significant (P less than 0.01) inhibition of the effects of PGE2 or PGD2 on CFU-E in murine bone marrow cultures. Butoxamine, a somewhat selective beta 2-adrenergic antagonist drug, also produced a significant inhibition of the effects of PGE2 on CFU-E in murine marrow cultures. These results indicate that the effects of beta-adrenergic blocking agents on prostaglandin-stimulated CFU-E are due to their membrane-stabilizing action rather than specific beta-adrenergic blockade.
Passive transport of K+ and Na+ in human red blood cells: sulfhydryl binding agents and furosemide
Authors: Wiater L. A. , Dunham P. B.
Published: December 2017

Passive transport pathways for K+ and Na+ were studied in fresh human red blood cells (pretreated with ouabain) by measuring unidirectional influxes. The effects of the sulfhydryl binding agents N-ethylmaleimide (NEM) and p-chloromercuribenzene sulfonate (p-CMBS) and the loop diuretic furosemide were studied. Influxes were measured at equimolar K+ and Na+ concentrations (50 mM) with both ions present and also in K+-free or Na+-free media. Some experiments were carried out in Cl--free media (with NO-3 as the substitute). NEM stimulated K+ influx twofold; the stimulation required Cl- but not Na+. NEM inhibited Na+ influx 20%. Furosemide inhibited both K+ and Na+ influxes. All of furosemide-inhibitable Na+ influx required the presence of K+. However 30% of furosemide-inhibitable K+ influx did not require Na+. All of furosemide-inhibitable K+ influx required Cl-. The ratio of Na+-dependent K+ influx to K+-dependent Na+ influx was 3:1. p-CMBS stimulated both Na+ and K+ influxes. K+ influx in p-CMBS cells required neither Na+ nor Cl-. Likewise p-CMBS-promoted Na+ influx did not require K+. These various results are consistent with two Cl--dependent pathways for K+ transport, one requiring Na+ [perhaps (Na + K + Cl) cotransport] and one independent of Na+ [perhaps (K + Cl) cotransport]. The pathways promoted by p-CMBS are probably independent of the apparent cotransport systems.
Tetrodotoxin-sensitive Na+ channels in isolated single cultured rat myocardial cells
Authors: Matsuki N. , Hermsmeyer K.
Published: December 2017

We studied the existence of tetrodotoxin (TTX)-sensitive fast Na+ channels in isolated single (verified by dye injection) myocardial cells compared with small multiple-cell groups from 1- to 3-day-old rat ventricles in cell culture. For single cells, average values were -63 mV maximum membrane potential, 32 mV overshoot, and 65 V/s maximum rate of rise of the action potentials (+Vmax). These values were comparable to values from groups of multiple (2-10) cells. TTX strongly depressed +Vmax dose dependently, with no difference between single and multiple cells. +Vmax was also decreased by lowering extracellular Na+ concentration but not by D 600 or lowering extracellular Ca2+ concentration. These results suggest that 1) isolated single cells possess TTX-sensitive fast Na+ channels, 2) culturing per se does not alter TTX sensitivity, and 3) TTX sensitivity is not modified by cell density.
NAD+-dependent ADP-ribosyltransferase in renal brush-border membranes
Authors: Kempson S. A. , Curthoys N. P.
Published: December 2017

Oxidized nicotinamide adenine dinucleotide (NAD+) in cytosol may interact with renal brush-border membranes (BBM) and inhibit BBM phosphate transport. The possible mechanism of interaction was investigated in the present study. Incubation of BBM with [adenine-3H]NAD+ led to acid-stable binding of 3H to the BBM, in contrast there was no binding of 14C when [carbonyl-14C]NAD+ was used. The data are consistent with an ADP-ribosylation mechanism involving transfer of ADP-ribose from NAD+ to BBM. This was confirmed by using [adenylate-32P]NAD+ and by the release of bound 32P in the form of 5'-[32P]AMP when the BBM were treated with snake venom phosphodiesterase. After gradient centrifugation of BBM the ADP-ribosyltransferase was recovered at the same density as known BBM enzymes, indicating that ADP-ribosyltransferase is an intrinsic BBM component and not a contaminant. These findings indicate that cytosolic NAD+ may be used for ADP-ribosylation of BBM proteins and that this may be a mechanism for regulating the BBM phosphate transport system.
Nocodazole and cytochalasin D induce tetraploidy in mammalian cells
Authors: Zieve G. W.
Published: December 2017

Nocodazole, a rapidly reversible inhibitor of microtubule assembly is useful for preparing mammalian cells synchronized at all stages of mitosis. When synchronized cells are allowed to progress through mitosis in the presence of cytochalasin D, the cleavage furrow is inhibited and dikaryon cells are formed. These cells become homogeneous populations of stable mononuclear tetraploid cells after the following cell division. This procedure is applicable to a wide range of mammalian cells in culture.
Protein secretion induced by isoproterenol or pentoxifylline in lacrimal gland: Ca2+ effects
Authors: Mauduit P. , Herman G. , Rossignol B.
Published: December 2017

In exorbital lacrimal glands, pentoxifylline (a methylxanthine) induces labeled protein secretion in a dose-related manner: the half-maximal and maximal stimulations are at 4 and 10 mM, respectively. In the presence of papaverine (10(-5) M), a phosphodiesterase inhibitor, labeled protein discharge is strongly stimulated by isoproterenol, via beta-adrenergic receptors: the maximal response is at 10(-6) M. l-Propranolol specifically inhibits the secretory stimulation to isoproterenol in a dose-related manner: for 5 X 10(-6) M isoproterenol in the presence of 10(-5) M papaverine, the half-maximal and maximal inhibitions are at 3 X 10(-7) and 10(-5) M, respectively. The beta-adrenergic response is mimicked by the adenosine 3',5'-cyclic monophosphate (cAMP) analogue dibutyryl cAMP (DBcAMP) at a 10(-3) M concentration. The time course of labeled protein secretion induced by pentoxifylline, DBcAMP, and isoproterenol shows a latency. In the presence or absence of extracellular calcium, pentoxifylline and isoproterenol immediately increase the cAMP intracellular level. Extracellular calcium omission increases the observed latency and also affects the maximal rate of protein secretion. As opposed to the cholinergic agonist, pentoxifylline has only a slight but sustained effect on 45Ca efflux, whereas isoproterenol has none. These data suggest that labeled protein secretion, such as that of peroxidase, can also be stimulated in rat exorbital lacrimal gland, through beta-adrenergic receptors; in the stimulation evoked by a beta-adrenergic agonist, DBcAMP, or methylxanthine, calcium could play a key role.