bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of sodium on beta-cell electrical activity
Authors: Ribalet B. , Beigelman P. M.
Published: December 2017

The present studies, designed to evaluate the contribution of Na+ to the mouse pancreatic beta-cell membrane potential, were performed utilizing intracellular microelectrodes. Complete removal of external sodium, in the presence of glucose, did not significantly affect spike peak potential. However, it caused a negative shift of the resting membrane potential, both in the presence and absence of glucose. After this initial hyperpolarization, the membrane gradually depolarized, the rate of depolarization being slower in the absence of glucose. This two-phase hyperpolarization-depolarization pattern remained when ouabain was added, both in the presence and absence of glucose. An increase of input resistance was associated with the slow depolarization. During this depolarization the maximum rate of rise (dV/dtmax) of the action potential (“spike”) decreased. There was no direct relationship between dV/dtmax and [Na]0. Readdition of low [Na]0 (14 mM) to a glucose medium reactivated the postburst hyperpolarization (PBH), even in the presence of ouabain. These observations indicate that there is a significant resting sodium permeability (PNa). However, the action potential (spike) is not generated by activation of a voltage-dependent (gated) sodium channel. The membrane depolarization after Na+ removal reflects concomitant inhibition of the Na+-K+ pump and decrease of potassium permeability (PK). The blockage of PBH in the absence of Na+ is not related to the inhibition of an oscillatory Na+-K+ pump but to the inactivation of a PK. Aside from its effect on the Na+-K+ pump, ouabain may stimulate PNa.
Effect of temperature, nutrients, calcium, and cAMP on motility of human spermatozoa
Authors: Gorus F. K. , Finsy R. , Pipeleers D. G.
Published: December 2017

The motility of human spermatozoa and its regulation were examined on cells isolated from other seminal components and purified into fractions of uniform progressive motility. The percent motile cells and estimates of their translational speed were determined by visual inspection, by stroboscopy, and by photon correlation spectroscopy; microcinematography and gradient centrifugation were occasionally used to clarify discrepancies. The motility of isolated spermatozoa could be maintained for periods up to 24 h at 4 or 37 degrees C; the presence of seminal fluid was not required and even provoked a reversible inhibition at 4 degrees C. Albumin facilitated cell movement between microscopic glass plates but had no effect on progressive motility per se, as evidenced by other techniques. During incubations of up to 2 h, progressive cell motility occurred independently of extracellular glucose and calcium but responded to variations in adenosine 3',5'-cyclic monophosphate and calcium. Dibutyryl cAMP increased forward motility, whereas ethyleneglycol-bis(beta-aminoethylether)-N,N'-tetraacetic acid reversibly immobilized the spermatozoa in a calcium-dependent manner; phosphodiesterase inhibition resulted in increased vibration of sperm heads without any effect on progressive motility. Longer incubation periods required the presence of extracellular nutrients. These experiments further demonstrate that several motility measuring techniques should be used in parallel to distinguish the various components of cell movement, to exclude aspecific effects, and to supplement the shortcomings of each individual technique. Such procedure could clarify the various discrepancies that have been reported so far and should lead to a better understanding of the regulation of human sperm motility.
Surfactant secretion: evidence that cholinergic stimulation of secretion is indirect
Authors: Massaro D. , Clerch L. , Massaro G. D.
Published: December 2017

There is strong evidence that cholinergic agents stimulate the secretion of surfactant in vivo and in the isolated perfused lung and that they do not stimulate surfactant secretion in isolated type 2 alveolar cells. These observations suggest that in multicellular systems the cholinergic effect is indirect. In the present work we have accrued the following support for this hypothesis. 1) Propranolol blocked the in vivo stimulation of disaturated phosphatidylcholine (DSPC) secretion by pilocarpine. 2) Bilateral adrenalectomy decreased by 50% the in vivo stimulation of DSPC secretion by pilocarpine. 3) Bilateral vagotomy did not block the increased secretion of DSPC produced in vivo by periodic deep inflations. 4) Pilocarpine (10(-7) M) stimulated DSPC secretion in the isolated perfused lung, and this effect was blocked by indomethacin as well as by atropine. We conclude that cholinergic stimulation of the secretion of surfactant in rats is indirect, i.e., cholinergic agonists do not stimulate the secretion of surfactant by acting directly on type 2 alveolar cells.
Placental amino acid uptake. VI. Regulation by intracellular substrate
Authors: Steel R. B. , Smith C. H. , Kelley L. K.
Published: December 2017

Amino acid uptake by human placental tissue is regulated by intracellular amino acids. alpha-Aminoisobutyric acid (AIB) uptake was reduced at intracellular AIB concentrations of 0.8 mM. The magnitude of reduction increased sharply between 1 and 3 mM and reached a maximum of 45% at 5 mM. Suppression was specific to the "A" system. It occurred only when both the amino acid used for preloading and that used as an uptake substrate were active with that system. In the "L" system, facilitation apparently occurs, and in the "ASC" system there is no apparent effect. The system specificity as well as other evidence indicated that suppression is caused by substrate present intracellularly rather than by dilution of extracellular substrate. Suppression was independent of inhibitors of protein synthesis and was not seen in membrane vesicles prepared from preloaded tissue, indicating that intracellular substrate interacts directly with the carrier (transinhibition) rather than altering its synthesis or degradation. The A system transinhibition has the potential to regulate syncytial uptake in vivo and limit variation due to changes in maternal plasma amino acid concentration.
Modulation of vasopressin action by reducing agents in Bufo marinus
Authors: Butkus D. E. , Schwartz J. H.
Published: December 2017

The effects of the reducing agent dithiothreitol (DTT) on vasopressin (AVP)-stimulated osmotic water flow and adenylate cyclase activity were studied in the urinary bladder of Bufo marinus. DTT produced concentration-dependent inhibition of the hydroosmotic water permeability response to 10 mU/ml AVP and 10 mM theophylline but did not inhibit the response to 10 mM adenosine 3',5'-cyclic monophosphate (cAMP). The inhibitory effects of DTT on AVP responsiveness were partially reversed by washing in DTT-free Ringer solution or by addition of oxidizing agents such as dehydroascorbic acid (DHA) or H2O2. The inhibitory effects of DTT were completely reversed by washing in DTT-free Ringer plus addition of DHA. In addition, the inhibitory effects of DTT on AVP-induced osmotic water flow were partially reversed by the GTP analogue 5'-guanylyl imidodiphosphate [Gpp(NH)p]. DTT also inhibited the adenylate cyclase response to AVP but did not alter the response to AVP plus Gpp(NH)p or the response to NaF. These observations suggest that the inhibitory effect of thiol compounds on AVP responsiveness may be modulated through alterations of a redox system distal to the hormone receptor but proximal to the catalytic subunit of adenylate cyclase. Inasmuch as Gpp(NH)p partially reversed the inhibitory effects of DTT on AVP-stimulated osmotic water permeability and prevented the inhibitory effect of DTT on AVP-stimulated adenylate cyclase, an effect on either GTPase or binding of GTP to the regulatory protein of adenylate cyclase is suggested by these observations.
Influence of hypothermia on intracellular pH during anoxia
Authors: Norwood W. I. , Norwood C. R.
Published: December 2017

Among the biochemical processes initiated by anoxia or ischemia that play a central role in cellular injury during deprivation is an alteration in cellular hydrogen ion concentration. In this study, the rate of exchange of intracellular hydrogen ion concentration in canine brain was compared with that in liver, using the 5,5-dimethyloxazolidine-2,4-dione method during anoxia at 37 and at 20 degrees C. The intracellular pH of brain decreased more rapidly than it did in liver anoxia at 37 degrees C. The intracellular pH of neither brain nor liver changed substantially during 30 min of anoxia at 20 degrees C. Although the ratio of tissue to plasma water did not change, the calculated extracellular-to-intracellular volume ratio increased during 30 min of anoxia at 37 but not at 20 degrees C.
Contractile properties of bundles of fiber segments from skeletal muscles
Authors: Faulkner J. A. , Claflin D. R. , McCully K. K. , Jones D. A.
Published: December 2017

Our purpose was to determine whether contractile properties of bundles of skeletal muscle fiber segments were significantly different from those of bundles of intact fibers. In frog muscles, the only difference between the contractile properties of fiber segments and intact fibers was a lower maximum velocity of shortening (Vo) for the fiber segments. In mammalian muscles, the contraction time (TPT), relaxation time (RT1/2), and maximum tetanus tension (Po) of bundles of fiber segments were not different from those of intact fibers, but the rate of tension development (dP/dt), twitch-to-tetanus ratio (Pt/Po) and Vo were lower. The lower dP/dt and Pt/Po resulted from increased compliance due to damaged sarcomeres near cut ends. Within 4-9 mm of a cut end, membrane potentials were less than control values, and sarcomeres lengthened during a fixed-end contraction. after the length of fiber segments was corrected for the exact portion that was not shortening, the Vo of fiber segments was not different from that of intact fibers. We conclude that valid estimates of contractile properties can be obtained from bundles of skeletal muscle fiber segments.
Effects of alterations in calcium levels on cat small intestinal slow waves
Authors: Mangel A. W. , Connor J. A. , Prosser C. L.
Published: December 2017

Intact segments of cat intestinal muscle and strips of isolated longitudinal muscle were treated with agents that reduce intracellular calcium concentration: incubation in 0-calcium saline, treatment with calcium conductance blockers, elevated extracellular magnesium concentration, or alkalinization with NH4Cl. These treatments reduced amplitude and frequency of slow waves in intact segments but only reduced frequency in isolated longitudinal muscle. The reduction in frequency was characterized by prolongation of the hyperpolarized phase of the slow waves. Treatments that would moderately increase intracellular calcium concentration, i.e., increasing external calcium to four times normal levels or lowering pH by CO2, increased slow-wave frequency. Increased frequency was associated with reduced amplitude and shortening of the hyperpolarized phase of the slow waves. Greater than four times normal calcium levels and intense spiking reduced slow-wave frequency. Chlorotetracycline fluorescence, an indicator of intracellular calcium concentration, showed fluctuations synchronous with slow waves. It is concluded that the reactions that pace the generation of slow waves are dependent on the level of intracellular calcium.
Effects of dantrolene and D2O on K+-stimulated respiration of skeletal muscle
Authors: Erlij D. , Shen W. K. , Reinach P. , Schoen H.
Published: December 2017

We have examined the effects of dantrolene and D2O on the K+-stimulated respiration in frog skeletal muscle. The threshold for K+ stimulation was around 10 mM extracellular potassium concentration ([K+]o). A further marked increase in respiration to levels about ten times the resting level was noted when [K+]o was between 15 and 20 mM. The increase was sustained for hours when [K+]o was less than 20 mM; however, with higher concentrations the stimulation consisted of an initial burst followed by a decline. Dantrolene shifted the relationship between [K+]o and peak increase in respiration toward higher [K+]o by about 10 mM; in addition it nearly completely blocked the sustained component of the increase. D2O, nearly abolished the K+-induced respiration. Neither agent shifted the relationship between [K+]o and membrane potential nor abolished the stimulation of respiration caused by caffeine. Dantrolene did not block the stimulation of Na+ efflux caused by 15 mM K+. The results with these agents are consistent with the proposal that K+-stimulated respiration is due to Ca2+ release into the cytoplasm. In addition, they provide evidence that the stimulated rate of Ca2+ release into the cytoplasm can remain at a persistently high level for hours provided [K+]o does not exceed 20 mM. We calculated that the level of this constant Ca2+ release is about 3.4 X 10(16) ions/(s.cm3).
Dome formation in primary cultured monolayers of alveolar epithelial cells
Authors: Goodman B. E. , Crandall E. D.
Published: December 2017

We have observed the formation of domes by type II alveolar epithelial cells harvested from rat lungs. The cells were harvested using elastase and grew to confluence in 3-4 days after plating on plastic. Numerous domes were observed in the monolayers 4-18 days after plating, with peak dome density occurring at days 6-9. When trypsin was used instead of elastase as the harvesting enzyme, many fewer domes were formed by the monolayers, with peak dome density observed at day 5 and no domes seen after 8 days. The life span of an individual dome was about 3-4 h. The presence of domes indicates an intact active transport function of the cells in the monolayer, which may represent an important mechanism for the maintenance of fluid-free air spaces and normal alveolar fluid balance in mammalian lungs in vivo.
Effect of disuse on sarcoplasmic reticulum in fast and slow skeletal muscle
Authors: Kim D. H. , Witzmann F. A. , Fitts R. H.
Published: December 2017

The effect of 6 wk of hindlimb immobilization on rat skeletal muscle sarcoplasmic reticulum (SR) was determined in the slow-twitch, type I soleus (SOL), the fast-twitch, type IIA deep region of the vastus lateralis (DVL), and the fast-twitch, type IIB superficial region of the vastus lateralis (SVL). Immobilization produced a significant decline in the Ca2+ uptake rate (Vmax) of SR vesicles from the slow SOL (0.930 +/- 0.116 to 0.365 +/- 0.071 mumol Ca2+ . mg-1 . min-1), while the SR Vmax increased in the fast SVL (2.763 +/- 0.133 to 5.209 +/- 0.687) and was unaltered in the DVL. Vesicles from the fast SVL and DVL also exhibited a higher total Ca2+ uptake capacity following immobilization. An evaluation of the time course of the immobilization-mediated effect revealed an increased Ca2+ uptake capacity in all three samples after 1 wk. In the SOL total Ca2+ uptake returned to control level after 2 wk, while in the fast-twitch muscles the higher capacities were maintained. The Ca2+-stimulated SR ATPase activity was not altered in any of the muscles studies, although the total SR ATPase activity increased twofold in the slow SOL.
Isometric contractile properties and velocity of shortening during avian myogenesis
Authors: Reiser P. J. , Stokes B. T. , Rall J. A.
Published: December 2017

Isometric twitch and tetanic contractile properties and velocity of unloaded shortening (V0) of whole avian posterior latissimus dorsi muscle (PLD) were examined between embryonic day 15 and the first 2 wk after hatching. The time to peak twitch force, time to half-relaxation of the twitch response, and time to half-peak tetanic force all change significantly during the final week in ovo but do not change during the first 2 wk ex ovo. Comparisons with previously published reports by others indicate that the twitch half-relaxation time at hatching is approximately the same as that of the adult PLD. The velocity of unloaded shortening increases 2.3-fold during the period studied. It has previously been shown by other that the velocity of shortening is well correlated with a muscle's myosin ATPase activity. Therefore, the observed changes in V0 suggest that the myosin ATPase activity of the avian PLD increases between embryonic day 15 and the first 2 wk posthatching, and this change could account, at least in part, for some of the changes in the isometric properties that were measured.
Hormonal stimulation of myoblast differentiation in the absence of DNA synthesis
Authors: Turo K. A. , Florini J. R.
Published: December 2017

The role of DNA synthesis in the final stages of muscle cell differentiation has been a subject of controversy for more than a decade. In an attempt to resolve disagreements over the necessity for a unique (or "quantal") mitosis just prior to the conversion of proliferating myoblasts to form postmitotic myotubes, we have studied the effects of insulin and somatomedin on the stimulation of myoblast differentiation with or without DNA synthesis. Under conditions in which at least 95% of [3H]thymidine incorporation was blocked by cytosine arabinoside, there was a 5- to 10-fold increase in the extent of differentiation (determined as fusion or creatine kinase elevation) on addition of insulin or multiplication-stimulating activity. The effect of the hormones was on myoblast differentiation, not enzyme induction; insulin did not cause any increase in creatine kinase when it was added to performed myotubes. These studies were done using two different cell types, Yaffe's L6 cell line and Japanese quail myoblasts in serum-free media; we obtained similar results in both. Our results are not compatible with the view that a quantal mitosis is required at a late stage of muscle cell differentiation.
Modulation of intracellular Na+ activity and cardiac force by norepinephrine and Ca2+
Authors: Lee C. O. , Vassalle M.
Published: December 2017

The actions of norepinephrine and high calcium on the electrical, mechanical, and intracellular sodium ion activities were studied in electrically driven canine cardiac Purkinje fibers under different conditions. It was found that norepinephrine and high calcium decrease intracellular sodium ion activity (aiNa). The exposure to either agent is followed by a transient decline of force that correlates with the lower aiNa. Inhibition of the Na+ -K+ pump by strophanthidin reduces or abolishes the decrease in aiNa by norepinephrine but not that by high calcium. It is concluded that norepinephrine and high calcium both decrease aiNa and thereby the contractile force but (unlike high calcium) norepinephrine acts through the stimulation of the Na+ -K+ pump.
Binding and antimineralocorticoid activities of spirolactones in toad bladder
Authors: Rossier B. C. , Claire M. , Rafestin-Oblin M. E. , Geering K. , Gaggeler H. P. , Corvol P.
Published: December 2017

The role of the soluble pool (cytoplasmic or cytosolic) of [3H]-aldosterone binding sites in the toad bladder was assessed by the use of two spirolactones, prorenone and spironolactone as a reference drug. Prorenone fulfills all the criteria for a specific competitive antagonist of aldosterone for its effect on Na+ transport. Compared with spironolactone (Ki approximately equal to 1 microM), prorenone was about eightfold less potent (Ki approximately equal to 8 microM). Competition for [3H]aldosterone binding sites by spironolactone and prorenone revealed an order of potency (spironolactone greater than prorenone) that corresponded to their antagonist activities in the Na+ transport assay. There was a linear correlation between the effects of the two spirolactones on the aldosterone-stimulated Na+ transport and their ability to displace [3H]aldosterone from its binding sites in the soluble pool. Finally [3H]prorenone binding sites were detected in the soluble pool but an insignificant number of antagonist-receptor complexes were found associated with the nuclear pool. Our study indicates that the aldosterone binding sites of the soluble pool are indeed mineralocorticoid receptors, which are probably the first intracellular mediators leading to an increased Na+ reabsorption.
Skin calcium-binding protein: distribution in other tissues
Authors: Pavlovitch J. H. , Didierjean L. , Rizk M. , Balsan S. , Saurat J. H.
Published: December 2017

The distribution of epidermal calcium-binding protein was examined in rat tissues using immunodiffusion and immunofluorescence techniques to investigate its possible physiological role. Epidermal calcium-binding protein was demonstrated in the basal proliferative cell layer of all Malpighian epithelia and related tissues (epidermis, sebaceous glands, cornea, esophagus, and vagina) as well as in ependyma of brain and in the epithelia of the lens. No immunoreactivity for epidermal calcium-binding protein was found in other tissues including dermis, muscle, cartilage, blood vessels, nerve tissue, liver, endocrine glands, urogenital tract, and intestinal and respiratory epithelium. The presence of a protein immunologically indistinguishable from epidermal calcium-binding protein in proliferative cells suggests that it may be involved in the control of calcium-dependent processes perhaps related to mechanical damage and continued proliferation.
Effects of detraining on enzymes of energy metabolism in individual human muscle fibers
Authors: Chi M. M. , Hintz C. S. , Coyle E. F. , Martin W. H. , Ivy J. L. , Nemeth P. M. , Holloszy J. O. , Lowry O. H.
Published: December 2017

Muscle biopsies were obtained from three cyclists and four runners at the end of 10-24 mo of intensive training and after intervals of detraining up to 12 wk. Control samples came from four untrained persons and four former athletes. Macro mixed fiber samples were assayed for lactate dehydrogenase, adenylate kinase, glycogen phosphorylase, citrate synthase, malate dehydrogenase, beta-hydroxyacyl-CoA dehydrogenase, succinate dehydrogenase, beta-hydroxybutyrate dehydrogenase, creatine kinase, hexokinase, 1-phosphofructokinase, fructosebisphosphatase, protein, and total creatine. In the case of three trained persons and two controls, the first six of the enzymes were also measured in individual fibers. Before detraining, enzymes of oxidative metabolism were substantially higher than in controls, and differences in levels between type I and type II fibers were smaller. During detraining, oxidative enzymes were decreased in both fiber types but the type II fibers did not fall to control levels even after 12 wk. Phosphorylase increased with detraining in both fiber types. The same is true for lactate dehydrogenase and adenylate kinase, except in the case of the type I fibers of one individual. Among the other six enzymes (measured in mixed fiber samples), only hexokinase was consistently affected (decreased) by detraining.
Mitogenic enhancement of purine base phosphoribosylation in Swiss mouse 3T3 cells
Authors: Becker M. A. , Dicker P. , Rozengurt E.
Published: December 2017

Increased uptake of guanosine and the purine bases, adenine, hypoxanthine, and guanine, occurs within minutes after addition of fresh serum or purified growth-promoting agents to density-arrested cultures of Swiss 3T3, Swiss 3T6, and tertiary mouse embryo fibroblasts. Enhancement of uptake is maximal by about 50 min, is potentiated by combinations of growth promoters, and involves a process distinguishable from that of the enhanced uridine uptake on the basis of time course, pattern of growth-factor responsiveness and the failure of uridine to inhibit purine uptake. Enhanced purine uptake in 3T3 cells results from stimulation of the phosphoribosylation of purine bases and consequent trapping of nucleotide derivatives rather than from increased rates of membrane transport of the purine compounds. Increased availability of 5-phosphoribosyl 1-pyrophosphate (PRPP), identifiable within 30 min of serum addition, provides a likely explanation for increased purine base phosphoribosylation. Mitogenic stimulation of purine uptake is unaffected by cycloheximide but is markedly reduced in sodium-free medium. Enhanced purine uptake does not clearly depend upon changes in concentrations of effectors of intracellular PRPP synthesis. Nevertheless, when the possibility of direct mitogenic activation of PRPP synthase was studied, no differences were found in activities of this enzyme in extracts from stimulated and unstimulated 3T3 cells and mouse embryo fibroblasts.
Release of chromaffin granular content on interaction with plasma membranes
Authors: Konings F. , Majchrowicz B. , De Potter W.
Published: December 2017

Cell-free interaction between bovine adrenal medullary plasma membranes and chromaffin granules results in the release of the granular content, a process specifically controlled by micromolar concentrations of calcium and therefore regarded as a putative model for exocytosis. The density gradient distribution of interacted organelles (1 min at 37 degrees C) was compared with the distribution of noninteracted material (kept at 0 degree C). Self-generating Percoll gradients, in which intact chromaffin granules were separated from plasma membranes and empty granule "ghosts," were used. The plasma membrane-induced release of the chromaffin granular matrix upon concomitant incubation was demonstrated by the disappearance of the "intact granule" peak accompanied by a proportional rise of the "granule ghost" peak. The degree of granule disappearance depended on the amount of plasma membranes added. The soluble content of the fraction that had disappeared was dispersed throughout the gradient. No shift of the granule peak toward intermediate densities was observed; this suggests that the loss of each granule's content is an all-or-none phenomenon rather than a partial release.
Hemolytic action of potassium salts on dog red blood cells
Authors: Parker J. C.
Published: December 2017

Recent demonstrations of chloride-associated passive potassium movements in red blood cells of humans, ducks, sheep, and toadfish prompted a reinvestigation of potassium permeability in dog red blood cells. Early observations of Davson (J. Physiol. London 101:265-283, 1942) had shown that replacement of chloride by nitrate and thiocyanate caused a greatly increased rate of potassium flux across the dog red cell membrane. This finding seemed at variance with results in other species in which chloride replacement caused a fall in potassium flux. The present data indicate that passive potassium movements in swollen dog red blood cells are chloride dependent and furosemide sensitive, as shown for the cells of other species. Davson's findings were demonstrated to be due to the inclusion of small quantities of calcium in the medium under circumstances that favored calcium entry into the cells, thus opening the calcium-activated potassium channel described by Gardos (Curr. Top. Membr. Transp. 10:217-277, 1978 and Nature London 279:248-250, 1979). Potassium movements through the latter channel were stimulated when chloride was replaced by more permeant anions, such as nitrate and thiocyanate, which also increased the rate of net potassium movements in valinomycin-treated cells.
Intracellular ion activities and Cl-transport mechanisms in bullfrog corneal epithelium
Authors: Reuss L. , Reinach P. , Weinman S. A. , Grady T. P.
Published: December 2017

Cell membrane potentials, cell membrane resistances, and intracellular ionic activities were measured in bullfrog corneal epithelium. Equivalent circuit analysis was performed by adding adenosine to the apical surface and assuming that only the apical membrane is initially affected. From single-ion substitutions in the apical bathing solution, the apical membrane was found to have a high Cl- permeability, a low K+ permeability, and an unmeasurably small Na+ permeability. Under control conditions intracellular Cl- activity (aCli) was 22 +/- 2 (SE) mM, intracellular Na+ activity (aNai) was 14 +/- 3 mM, and intracellular K+ activity (aKi) was 106 +/- 5 mM. The electrical potential differences across apical and basolateral membranes were about 50 and 67 mV, respectively, both cell negative. aCli and aKi are higher, whereas aNai is much lower than predicted for equilibrium distribution. Inasmuch as Cl- is transported from the basolateral (stromal) to the apical (tear) side, basolateral entry of this anion is uphill and apical exit is downhill. Basolateral entry is Na+ dependent, as evidenced by a fall of aCli to near-equilibrium values after basolateral Na+ removal. The electrochemical gradient for Cl- efflux across the apical membrane is large enough to account for Cl- transport by electrodiffusion only. Na+ removal from the basolateral solution causes a reversible decrease of apical membrane Cl- permeability. The results support the hypothesis that net transepithelial Cl- transport results from coupled NaCl entry (or an equivalent process) at the basolateral membrane and electrodiffusional Cl- exit at the apical membrane.
Levels of enzymes of energy metabolism are controlled by activity of cultured rat myotubes
Authors: Lawrence J. C. , Salsgiver W. J.
Published: December 2017

We have investigated the effects of inhibiting the spontaneous activity of cultured rat myotubes on several representative enzymes of glycolytic and oxidative metabolism. The results presented demonstrate that contractile activity in the absence of nerves can regulate the amounts of these enzymes and indicate that muscle activity may partially control development of the metabolic types of muscle fibers. Control muscle cells have relatively high levels of glycolytic enzymes and low oxidative enzymes and metabolically most closely resemble fast glycolytic fibers. The divalent cation ionophore A23187 caused enzyme levels of the cultured cells to change towards those found in tonically contracting skeletal muscle fibers in vivo. The evidence presented suggests that calcium may mediate certain of the effects associated with muscle contraction on enzymes of energy metabolism.
Evidence for basolateral membrane potassium conductance in canine tracheal epithelium
Authors: Welsh M. J.
Published: December 2017

The ionic dependence of the basolateral membrane conductance in canine tracheal epithelium was investigated using intracellular microelectrode techniques. Increasing the K+ concentration in the submucosal bathing solution depolarized the electrical potential difference across the basolateral membrane; neither alteration of the submucosal Na+ concentration nor the mucosal K+ concentration had a significant effect on the cellular electrical potential profile. An increase in the K+ concentration in the submucosal bathing solution also decreased the net rate of Cl-secretion. Addition of ouabain (10(-4) M) to the submucosal bathing solution decreased the short-circuit current and depolarized the intracellular voltage without altering transepithelial resistance or the cell membrane resistance ratio, suggesting that basolateral resistance was unchanged. These findings, together with the previous observation that there is no appreciable basolateral Cl- conductance, indicate that a K+ conductance accounts for the predominance of the electrical conductance at the basolateral membrane. The results also indicate that the basolateral membrane K+ conductance plays a critical role in the generation of the negative intracellular voltage that drives Cl- exit across the apical membrane and thus supports Cl- secretion.
Beta-adrenergic modulation of mucin secretion in cat trachea
Authors: Liedtke C. M. , Rudolph S. A. , Boat T. F.
Published: December 2017

The mechanism of beta-adrenergic regulation of mucin secretion was investigated in cat trachea in vitro. beta-Adrenergic agonists increased the release of [35S]SO4-radiolabeled mucin and mucosa-submucosal adenosine 3',5'-cyclic monophosphate (cAMP) levels in a dose- and time-dependent manner. The relative potencies and efficacies of l-isoproterenol, l-epinephrine, l-norepinephrine, terbutaline, and dobutamine for physiological and biochemical events were similar. The effect of these agonists were blocked by d-l-propranolol. 3-Isobutyl-l-methylxanthine (IBMX) and 8-bromo-cAMP mimicked the effects of the agonists on mucin release. IBMX increased cAMP levels and potentiated the increase in cAMP levels effected by beta-adrenergic agonists. The half-maximal effects of l-isoproterenol on cAMP levels and mucin release were attained at 1.6 and 8.8 min, respectively. Three major mucosa-submucosal proteins of apparent molecular weights of 49,000, 54,000, and 59,000 daltons displayed reduced binding of the photoaffinity label 8-N3-[32P]cAMP when endogenous cAMP levels were increased with l-isoproterenol and/or IBMX. The first two proteins correspond in electrophoretic mobility to the regulatory subunits of type I and type II cAMP-dependent protein kinases, respectively. The 59,000-dalton cAMP binding protein may be the phosphorylated form of the regulatory subunit of type II cAMP-dependent protein kinase. These data are consistent with the hypothesis that beta-adrenergic modulation of tracheal mucin release is mediated by cAMP and suggest that activation of cAMP-dependent protein kinases may be involved in the neurohormonal effects.
Role of membrane fusion in CO2 stimulation of proton secretion by turtle bladder
Authors: Stetson D. L. , Steinmetz P. R.
Published: December 2017

The changes in cell structure produced during stimulation of proton secretion by CO2 in turtle bladder were examined using ultrastructural morphometric methods. One hour after CO2 addition, the area of the luminal membrane of the carbonic anhydrase-containing (CA) cell population was increased 2.5-fold and the volume percent of electron-lucent cytoplasmic vesicles in these CA cells was decreased by 61%. No changes were observed in the epithelial granular cells. These results suggest that during CO2 stimulation the vesicles fuse with the luminal membrane. CO2 stimulation of proton secretion is inhibited by the cytoskeleton-disrupting drugs colchicine and cytochalasin B and by 99% deuterium oxide as the Ringer solvent. Deuterium oxide also inhibits the decrease in cytoplasmic vesicles. Thus stimulation of proton secretion by turtle bladder CA cells depends to a large extent on vesicle fusion and the resultant increase in luminal surface area.
Complex regulation of fibronectin synthesis by cells in culture
Authors: Senger D. R. , Destree A. T. , Hynes R. O.
Published: December 2017

We have studied the biosynthesis of fibronectin by NIL8 hamster embryo cells in various stages of growth. Pulse labeling with [35S]methionine, immunoprecipitation, electrophoresis, and autoradiography were employed to compare fibronectin synthesis by cells in subconfluent monolayers, confluent monolayers, and “aged” postquiescent monolayers. We have determined that fibronectin synthesis is proportionally low while cells are subconfluent, rises to maximal levels at confluence but just prior to quiescence, and then declines with increasing culture age in quiescent cultures. Furthermore, when cells are stimulated to grow, the effects on rates of fibronectin synthesis depend on the prior history of the cells; freshly confluent cells stimulated to grow show reductions in rate of fibronectin synthesis, whereas aged postquiescent cultures show increases in this rate. These results are in contrast to those on the rates of synthesis of other proteins (including the precursor to the C3 component of complement), which strictly correlate with quiescence and do not decline significantly with culture age postconfluence. We have also determined that the microheterogeneity of pulse-labeled fibronectin differs between exponentially growing and quiescent cells and that it becomes less heterogeneous once cells are quiescent. We conclude that the microheterogeneity of pulse-labeled fibronectin and the proportionate amount of total fibronectin synthesized both depend on growth state prior to the entry of cells into quiescence and that they depend on culture age thereafter.
A novel effect of amiloride on H+-dependent Na+ transport
Authors: Dubinsky W. P. , Frizzell R. A.
Published: December 2017

The diuretic amiloride stimulates the rate of decay of a proton gradient in both native ileal brush border membrane vesicles and artificial phospholipid vesicles. The kinetics of the rate of decay of the pH gradient were studied using acridine orange in membrane vesicles equilibrated at pH 5.5 and assayed at pH 8.0. Amiloride stimulated the decay of the proton gradient in a dose-dependent fashion. In parallel experiments employing artificial phospholipid vesicles and the monovalent cation-H+ exchanger nigericin, amiloride inhibited both the initial rate and extent of nigericin-mediated 22Na+ accumulation. It is concluded from these studies in both native and artificial membrane vesicles that amiloride can act as permeant weak base thus dissipating the pH gradient necessary for Na+ accumulation via a Na+-H+ exchanger.
Actin filaments and vasopressin-stimulated water flow in toad urinary bladder
Authors: Pearl M. , Taylor A.
Published: December 2017

Vasopressin increases the water permeability of the apical membrane of the granular epithelial cells of the toad urinary bladder. Cytochalasin B inhibits this action of the hormone, indicating that microfilaments may play a role in the water permeability response. We have extended previous functional studies with cytochalasin B and have demonstrated that dihydrocytochalasin B, a more specific inhibitor of actin filament elongation, similarly diminishes the hydrosmotic response to vasopressin. Biochemical studies of isolated epithelial cells indicate that an actin-like protein accounts for about 10% of the soluble protein of the epithelium. Morphological studies of whole toad bladders incubated with heavy meromyosin conclusively demonstrate that actin is a component of the epithelial cells and that actin-containing filaments are associated with both plasma membranes and cytoplasmic organelle membranes. Taken together, these findings provide strong, albeit indirect, evidence that actin microfilaments play a functional role in the hormone-induced increase in water permeability in the toad urinary bladder.
beta-Adrenergic catecholamine-dependent properties of rat myometrium primary cultures
Authors: Fortier M. , Chase D. , Korenman S. G. , Krall J. F.
Published: December 2017

We have used cultured trypsin-collagenase-dispersed cells from uteri of 21-day-old rats to investigate the mechanism of control of uterine motility by the beta-adrenergic receptor. After 5 to 7 days in RPMI 1640 the cells started to assume some of the morphological characteristics of smooth muscle cells. When cultures were incubated with 45Ca2+ for 3 h then washed free of isotope and incubated in medium with unlabeled Ca2+, efflux from the prelabeled intracellular pools was linear for up to 60 min. The potent beta-adrenergic agonist isoproterenol had a rapid effect on the rate of efflux and increased it almost sevenfold. Isoproterenol's effect was blocked by propranolol and could be duplicated by the addition of 8-bromo-adenosine 3',5'-cyclic monophosphate or cholera toxin. The cultured myometrial cells had adenylate cyclase properties similar to those of intact muscle strips when these were determined by the conversion of radioactive substrate (alpha-32P-ATP) to 32P-cAMP using a broken-cell preparation. Adenylate cyclase was sensitive to stimulation by GTP and by isoproterenol in the presence but not in the absence of GTP. Adenylate cyclase was also sensitive to stimulation by Ca2+ in the absence of GTP. We conclude that the primary cultures had the properties expected of smooth muscle cells including beta-adrenergic receptors that were coupled to a physiologically important function, Ca2+ flux. The beta-adrenergic receptor's effect on Ca2+ flux was cAMP mediated, and the divalent cation may also regulate its rate of flux by an effect on Ca2+-sensitive cAMP production.
Influence of organic acids on intracellular pH
Authors: de Hemptinne A. , Marrannes R. , Vanheel B.
Published: December 2017

By use of double-barreled pH-sensitive microelectrodes, intracellular pH was measured in isolated sheep cardiac Purkinje strands. After equimolar substitution of 20 mmol/l Cl- by several organic anions at constant extracellular pH 6.8, the rate of induced intracellular acidification was measured. For many organic acids tested, a relation was found between the rate of intracellular acidification and the product of their dissociation constant (pK'a) and diisopropylether-to-water partition ratio (p'). L-Lactate and pyruvate, and also cyanoacetate and alpha-ketobutyrate, caused faster acidifications than anticipated from their pK'a and p'. The rate of intracellular acidification, induced by L-lactate and pyruvate, was markedly depressed in the presence of 4 mmol/l alpha-cyano-4-hydroxycinnamate, a known inhibitor of the carrier-mediated pyruvate transport. The drug also had an effect on the acidification produced by cyanoacetate, alpha-ketobutyrate, glycolate, alpha-hydroxybutyrate, and alpha-chloropropionate, but not on that produced by propionate and acetate. L-Lactate caused a faster acidification than D-lactate. Our results suggest the existence of a facilitated diffusion for L-lactate, pyruvate, and some other organic acids in sheep Purkinje cells.
Cholinergic potentiation of isoproterenol-induced cAMP level in sweat gland
Authors: Sato K. , Sato F.
Published: December 2017

Although methacholine (MCh) and a Ca2+ ionophore A23187 do not enhance the tissue adenosine 3',5'-cyclic monophosphate (cAMP) level by themselves, they markedly potentiate isoproterenol (ISO)-induced tissue cAMP accumulation in isolated simian eccrine sweat glands in a dose-dependent manner. The agonist concentration producing 50% of the maximal response of such a potentiated cAMP accumulation was 2.1 X 10(-7) M for MCh, 2.5 X 10(-7) M for ISO, and 2.9 X 10(-6) M for A23187. Unlike cAMP accumulation induced by ISO alone, MCh-stimulated ISO-induced cAMP accumulation is dependent on extracellular Ca2+. MCh- plus ISO-induced cAMP level is tripled by 10(-2) M theophylline (TH), and while the ISO-induced cAMP level was also elevated by TH, it was not enhanced to the level of ISO-plus MCh-induced cAMP accumulation, indicating that phosphodiesterase inhibition is not the major mechanism for the augmentative effect of MCh. The augmentative effect of MCh was seen only in the secretory portion, whereas that of A23187 was seen in both the secretory portion and the duct. The data suggest that MCh-induced augmentation of ISO-stimulated cAMP accumulation is due to increased cAMP formation, not decreased cAMP degradation, and that it may be mediated by an elevated intracellular Ca2+.
Stereospecific glucose transport across motor nerve terminal membrane: an electrophysiological study
Authors: Shimoni Y. , Rahamimoff R.
Published: December 2017

Spontaneous transmitter release at the neuromuscular junction of the frog and rat was monitored during exposures to hyperosmotic solutions containing different sugars. Raising the osmolarity of the medium with D-glucose causes a marked, but transient, increase in the frequency of miniature end-plate potentials (MEPPs): after the initial elevation in frequency there is a subsequent decline towards the control levels, in spite of a continuous perfusion with the hyperosmotic solution. This decline occurs more rapidly in the frog. Two nonmetabolized analogues of glucose, 2-deoxy-D-glucose and 3-O-methylglucose, cause a transient hyperosmotic increase in MEPP frequency, which is very similar to the effect of D-glucose. The elevation of MEPP frequency with hyperosmotic glucose is stereospecific. Hyperosmotic solutions of L-glucose cause a sustained increase in transmitter release in the rat and frog. Insulin dramatically reduces the response of the frog nerve terminal to hyperosmotic D-glucose. Phenolphthalein, a glucose transport blocker, reduces or eliminates the secondary decline in MEPP frequency. It is suggested that the transient nature of the response to hyperosmotic solutions reflects the penetration of the hyperosmotic agent into the nerve terminal. The rate of decline of the MEPP frequency presumably indicates the rate of transport, which determines the rate of osmotic equilibration. This rate can then serve as an index of the relative permeability of the functioning presynaptic membrane to different sugars.
Ca2+-dependent slow action potentials in normal and dystrophic mouse skeletal muscle
Authors: Kerr L. M. , Sperelakis N.
Published: December 2017

Slowly rising action potentials (APs), previously described in amphibian skeletal muscle, were examined in skeletal muscle of normal and dystrophic mice (129/ReJ strain). A standard two-microelectrode recording technique was used. Muscles were bathed in a solution that was Cl- free (methanesulfonate substituted), high in K+ (20 mM), and contained 15 mM tetraethylammonium. The slow APs were elicited under conditions in which the fast Na+ channels were voltage inactivated (by partial depolarization) and in which the external Na+ concentration was only 10 mM. Increases in external Ca2+ concentration produced increases in slow AP amplitude and duration. Mn2+ (4 mM), La3+ (4 mM), and detubulation with osmotic shock blocked the slow APs. When slow APs were generated at 30-s intervals, their amplitude stayed constant. When they were generated at 15-s intervals, their amplitude decreased progressively and then fell to zero by the 11th stimulus. The Ca antagonists verapamil (10(-5) M) and bepridil (10(-5) M) caused this decrease in amplitude to occur more quickly. Voltage inactivation of the slow APs occurred between -45 and -10 mV. Slow APs recorded from dystrophic muscle fibers were decreased in amplitude and duration compared with those in normal fibers, and there was a reduced incidence of occurrence; 96% of the fibers in normal muscle exhibited slow APs compared with only 46% of dystrophic muscle fibers. In summary, slow Ca2+ APs in mammalian muscle are similar to those in cardiac and amphibian skeletal muscle, and these slow APs are depressed in dystrophic skeletal muscle.
Thiol-dependent passive K+-Cl- transport in sheep red blood cells. V. Dependence on metabolism
Authors: Lauf P. K.
Published: December 2017

Measurement of ouabain-insensitive K+ efflux and Rb+ influx in low-K+ sheep red blood cells with artificially altered cellular ATP levels revealed that Cl--dependent K+ transport was activated by N-ethylmaleimide (NEM) in cells with ATP concentrations above 0.5 mM. Depletion of ATP by starvation at 37 degrees C either in glucose-free media for 16 h or in 2-deoxy-D-glucose-containing media for 4 h completely abolished the response of K+-Cl-transport to NEM but did not reduce the basal Cl--dependent K+ flux. On repletion of cellular ATP by a second incubation in media containing glucose, inosine, and inorganic phosphate, the NEM-stimulated K+ (Rb+) flux reappeared. The magnitude of flux reactivation varied directly and monotonically with the ATP level. The data constitute the first unequivocal evidence for a reversible ATP dependence of thiol groups functionally involved in the thiol-dependent K+-Cl- transporter, suggesting that thiol-dependent and volume-sensitive K+-Cl- transport systems are operationally different.