bluuebunny/crossref_metadata_2025 huggingface dataset:
Voltage-dependent Ca2+ channel and Na+ channel in frog taste cells
Authors: Kashiwayanagi M. , Miyake M. , Kurihara K.
Published: December 2017

Frog taste cells were hyperpolarized by injecting an inward current pulse, and regenerative anode-break potentials were observed at the termination of the current pulse. The results obtained are as follows. 1) The magnitude of the anode-break potentials increased with the extent of hyperpolarization of taste cells and reached a saturation level around -200 mV. 2) The magnitudes of the anode-break potentials observed in 80 different taste cells hyperpolarized to about -200 mV were distributed widely from cell to cell. The average magnitude was 39 mV. 3) The anode-break potentials were recorded after the lingual artery was perfused with artificial solutions containing various channel blockers. The results indicated that the anode-break potentials are composed of Na+ and Ca2+ components. 4) The slope of the current-voltage relation obtained with cells hyperpolarized to 100 mV was appreciably decreased above -50 mV by application of tetrodotoxin to the perfusing solution. Discussion was made on possible roles of the voltage-dependent Na+ and Ca2+ channels in the electrotonic spreading of the depolarization at the receptor membranes to the synaptic area and in releasing a chemical transmitter.
Linear range of Na+ pump in sciatic nerve of frog
Authors: Brink F.
Published: December 2017

The Na+-activated utilization of O2 by frog sciatic nerve is a linear function of the internal concentration of Na+ in preparations deprived of external Ca2+. It is assumed that this component of O2 uptake is a measure of the active extrusion of Na+ by the Na+ pump because it is suppressed by ouabain. However, no tendency toward saturation, expected in an enzymatic process with a high affinity for Na+, was evident. Similar linearity has been reported in other neuronal preparations in experiments measuring 22Na+ efflux directly. The slope of the linear relation between Na+-activated O2 uptake and the concentration of internal Na+ is independent of the external concentration of Na+, but it decreases when the external concentration of K+ is changed from 10 to 4.8 mM.
Active electrogenic mechanisms for alkali and acid transport in turtle bladders
Authors: Satake N. , Durham J. H. , Ehrenspeck G. , Brodsky W. A.
Published: December 2017

Immediately after mounting in the Ussing chamber between choline bicarbonate Ringer solutions devoid of exogenous Na and Cl, the serosal fluid is electronegative to the luminal fluid in bladders from postabsorptive and acidotic turtles; and electropositive in bladders from alkalotic turtles. In bladders from postprandial turtles, the electrical orientation, initially serosal positive, reverses to serosal negative. Serosal additions of 3-isobutyl-1-methylxanthine (IBMX) and adenosine 3',5'-cyclic monophosphate (cAMP) produce no changes in the negative short-circuiting current (Isc) of acidotic turtles but induce large positively-directed increases of Isc in bladders from other turtle groups. With IBMX and cAMP in the (HCO3 + CO2)-rich serosal fluid at pH 7.2 and with luminal pH maintained at 4.0-5.0, the rate at which titratable alkali enters the luminal fluid is electrochemically equal to the positive Isc; and this increased positive Isc is the same as that in the absence of transepithelial gradients. The effects of acetazolamide and 4-acetamido-4-isothiocyanostilbene-2,2'-disulfonic acid on positive and negative Isc are presented. It is concluded that isolated bladders from alkalotic, postprandial or postabsorptive turtles, but not those from acidotic turtles, possess an active electrogenic mechanism for a Na-independent Cl-independent secretion of bicarbonate. This transport process is accelerated by phosphodiesterase inhibitors (IBMX) and cAMP or its eight substituted derivatives.
Cerebral glucose utilization in awake unstressed rats
Authors: Bryan R. M. , Hawkins R. A. , Mans A. M. , Davis D. W. , Page R. B.
Published: December 2017

Regional cerebral glucose utilization (rCMRGlc) was measured in awake unstressed rats by use of [2-14C]glucose autoradiography. Rats prepared by chronic catheterization of the jugular and epigastric veins were placed in specially designed chambers after surgery. The catheters were passed through the top of the chambers. Seven days after surgery, plasma glucose was normal (7.5 mM), plasma ketone bodies were low (0.13 mM), and body weight was maintained. Plasma epinephrine was 31 pg/ml; approximately 3% of the level for stressed (immobilized) rats. Injections and blood sampling through the catheters did not produce changes in heart rate, blood pressure, plasma catecholamines, or blood metabolites. Visual observations did not reveal any signs that the rats were aware of the [2-14C]glucose injections and the subsequent blood sampling. The rCMRGlc in stressed rats was significantly greater than in unstressed rats. This method provides a model for physiological studies in unstressed unanesthetized rats.
Effects of calcium on membrane potential and sodium influx in barnacle muscle fibers
Authors: Sheu S. S. , Blaustein M. P.
Published: December 2017

The influence of internal and external Ca2+ on membrane potential and 22Na influx were tested in internally perfused giant barnacle muscle fibers. The fibers depolarized by about 2-3 mV, and Na+ influx increased when external Ca2+ was removed. These effects were inhibited and reversed by adding 2 mM La3+ externally but not by tetrodotoxin (TTX). Ca2+ channel blockers did not prevent the depolarization. Increasing internal free Ca2+ ([Ca2+]i) from 10(-7) to 10(-5) M also stimulated Na+ influx and depolarized the fibers by a few millivolts. Neither external La3+ nor TTX prevented the effects of raising [Ca2+]i; however, internal tetrabutylammonium ions depolarized the fibers and attenuated the internal Ca2+-dependent effects. These data are consistent with the idea that removal of external Ca2+ activates a La3+-sensitive channel that is permeable to Na+; raising [Ca2+]i activates a La2+-insensitive, Na+-permeable channel that may be similar to the internal Ca2+-activated nonselective cation channels observed in cardiac muscle. The results demonstrate that all Na+ (and Ca2+) fluxes that do not contribute to Na-Ca exchange must be carefully identified before the exchange stoichiometry can be determined from Na+ and Ca2+ flux measurements.
Excitation-contraction coupling without Ca2+ action potentials in small intestine
Authors: Sanders K. M.
Published: December 2017

Sensitive mechanical and intracellular electrical recordings showed that phasic contractions occurred in response to electrical slow waves in the absence of Ca2+ action potentials. Drugs that either enhanced or depressed slow waves were used to study the relationship between slow-wave amplitude and the amplitude of the phasic contractions. Acetylcholine (Ach) (10(-8) to 3 X 10(-7) M) increased slow waves and contractions without causing action potentials. When ACh was raised to 10(-6) M, action potentials were elicited and accompanying contractions increased in amplitude by at least a factor of five. The Ca2+ channel blocker, Mn2+ (0.5 mM), decreased slow-wave amplitude and the associated phasic contractions. These data agree with a previous study (12), suggesting that an oscillation in intracellular Ca2+ occurs during each slow-wave cycle. The present study suggests that the increase in intracellular Ca2+ during the slow wave is sufficient to activate the contractile apparatus.
Inhibition of prostaglandin biosynthesis in renal (MDCK) cells by cAMP
Authors: Hassid A.
Published: December 2017

Cultured renal tubular cells (MDCK) have many of the biological properties of renal medullary tubular epithelial cells, including the ability to synthesize prostaglandin E2 (PGE2) as the major arachidonate metabolite. The hypothesis that adenosine 3',5'-cyclic monophosphate (cAMP) regulates prostaglandin synthesis in these cells was investigated by using cAMP, two degradation-resistant cAMP analogues [8-bromo-cAMP (8-BrcAMP) and N6,O2'-dibutyryl cAMP (DBcAMP)], and a phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine (IBMX). These agents inhibited basal-, calcium ionophore (A23187)-, or bradykinin-stimulated PGE2 biosynthesis by MDCK cells. The observed inhibition was dose- and time-dependent and could be reversed after 30 min of incubation in the absence of inhibitor. IBMX dose-dependently increased intracellular and extracellular cAMP levels by severalfold, suggesting that it was inhibiting prostaglandin biosynthesis by increasing cellular cAMP levels. Vasopressin, which stimulated cAMP levels by less than two-fold, did not inhibit prostaglandin synthesis. 8-BrcAMP and N6,O2'-DBcAMP inhibited A23187- or bradykinin-stimulated release of [3H]arachidonate from prelabeled cells, suggesting that cAMP inhibited acylhydrolase activity. Moreover, 8-BrcAMP also inhibited the conversion of exogenous arachidonate to PGE2 in intact cells and in a subcellular fraction containing prostaglandin synthetase activity, suggesting that cAMP inhibited cyclooxygenase and/or PGE2 isomerase activity. cAMP thus appears to regulate prostaglandin biosynthesis in MDCK cells by modulating the activity of two or more of the enzymes involved in the biosynthetic process.
Reversible carbon dioxide-induced inhibition of dye coupling in Necturus gallbladder
Authors: Jarrell J. A.
Published: December 2017

The cells of Necturus gallbladder epithelium are electrically coupled. This work used intracellular injection of the fluorescent dye Lucifer yellow to demonstrate that these cells are also dye coupled and that this coupling is rapidly and reversibly inhibited by high concentrations of carbon dioxide. Dye coupling is also inhibited by the calcium ionophore A23187.
Single K+-channel current measurements from brain synaptosomes in lipid bilayers
Authors: Nelson M. T. , Roudna M. , Bamberg E.
Published: December 2017

Ion channels from a rat brain preparation enriched in presynaptic nerve terminals (synaptosomes) were incorporated into planar lipid bilayers. Experiments examined macroscopic (channel-ensemble) currents as well as single-channel currents. Four single-channel conductances (ranging from 10 to 40 pS) were usually observed, each with distinct kinetic properties. All the observed channels selected for K+ over Cl-. These K+ channels may contribute to the resting K+ conductance of brain nerve terminals. Furthermore, this report demonstrates that the properties of ion channels from mammalian brain can be studied in planar lipid bilayers and suggests that this system can be readily extended to many additional investigations on the electrical properties of brain membranes.
Origin of the decline in pancreatic amylase secretion with time in vitro
Authors: Ho J. J. , Rothman S. S.
Published: December 2017

The release of alpha-amylase (EC 3.2.1.1) into the medium bathing strips of rabbit pancreas decreased gradually with time in vitro so that after 3 h of incubation the secretory rate was only 20% of that seen initially. This decrease was not the result of an intrinsic loss of cellular responsiveness or viability, because a cholinergic agonist was still able to augment amylase secretion from the tissue after this incubation period to the same, or greater, degree than from “fresh” tissue. Rather, the inhibition of secretion was apparently due to the release of a nondialyzable substance from the tissue, which appears to be amylase itself. The observations that led to this conclusion are 1) replacing the bathing medium every 30 min eliminated the time-dependent decrease in enzyme secretion, 2) this replacement was ineffective if the tissue was separated from the medium by a dialysis membrane, and 3) adding an excess of the amylase substrate glycogen to the medium also prevented the falloff in amylase secretion, the effect of glycogen presumably being due to the formation of an enzyme-substrate complex in the medium. These observations are consistent with other observations that demonstrate that the rate of enzyme secretion by the pancreas depends on the rate of removal of product from the site of its secretion [Isenman and Rothman, Proc. Natl. Acad. Sci. USA 74: 4068-4072, 1977; Isenman and Rothman, Science 204: 1212-1215, 1979; Ho and Rothman, Am. J. Physiol. 242 (Gastrointest. Liver Physiol. 5): G32-G39, 1982].
Demonstration of an electrogenic Na+-K+ pump in mouse spleen macrophages
Authors: Gallin E. K. , Livengood D. R.
Published: December 2017

The effect of ouabain on the membrane potential of cultured mouse spleen macrophages was examined. Ouabain (10(-3) M) induced a membrane depolarization (6.6 mV) in 18 of 19 cells studied that occurred within several minutes after exposure and was not associated with significant changes in current-voltage relationships. In related studies, cells were placed in K+-free medium in the cold for 2 h to block pump activity. Subsequent exposure to K+ at 37 degrees C resulted in a membrane hyperpolarization. However, addition of K+ also enhanced inward rectification. To differentiate between the effect of K+ on the Na+-K+ pump and its action on inward rectification, two types of experiments were done. Studies performed in the presence of barium, which blocks inward rectification, demonstrated a K+-induced hyperpolarization with no changes in rectification. Additional studies examined the effects of rubidium on Na+-loaded cells. Rubidium, which blocks inward rectification but substitutes for K+ in activating the Na+-K+ pump, induced membrane hyperpolarizations that were reversed by addition of ouabain. These data indicate that macrophages exhibit an electrogenic Na+-K+ pump, which probably contributes to the resting membrane potential under steady-state conditions and can be activated under conditions designed to Na+ load the cells. In addition, they demonstrate that increasing extracellular K+ enhances inward rectification in macrophages.
Cytoplasmic involvement in ADH-mediated osmosis across toad urinary bladder
Authors: DiBona D. R.
Published: December 2017

Several lines of investigation have suggested that antidiuretic hormone (ADH) may have direct effects on the cytoskeletal organization of granular epithelial cells in the toad urinary bladder. To some extent, these effects are in concert with the well-established action of ADH on the hydraulic permeability of the mucosal plasma membrane, but it appears that other conformational adjustments (largely cytoplasmic) may be of comparable importance. The thrust of this review is that the hormone brings about a general restructuring of the granular cells so that the epithelium as a whole may function efficiently as an osmotic pathway. Details of cytoskeletal changes are far from clear as yet, but interference with or modulation of these particular effects infer that cytoplasmic organization is the seat of feedback control of osmotic flow rate, the basis for viability in the presence of dramatic cytosolic dilution and a major factor in the observed disparity in osmotic and diffusional permeability coefficients. In the interest of stimulating new thoughts and experiments in this area, a number of preliminary findings have been freely cited.
Tension and ATPase rate in steady-state contractions of rabbit soleus fiber segments
Authors: Krasner B. H. , Kushmerick M. J.
Published: December 2017

Steady-state isometric tension and ATPase were studied in hyperpermeable segments of single muscle fibers from rabbit soleus muscle at 22 degrees C. The ATPase activity was due to actomyosin. The ratio of fiber ATPase to tension was used as an index of steady-state cross-bridge kinetics. Increasing the calcium ion concentration from pCa 8 to pCa 5 activated both tension and ATPase. The maximal tension was 1.35 +/- 0.07 kg/cm2. The maximal ATPase was 1.05 +/- 0.13 mumol X g-1. s-1 at pCa 5.2. ATPase activity increased with tension, such that the ratio of ATPase to tension remained constant at all calcium concentrations. In the absence of calcium, increasing the concentration of MgATP from 1 to 7 X 10(-7) M increased tension from zero to a maximum of 0.46 +/- 0.03 kg/cm2. Increasing MgATP concentration further to 1 X 10(-6) M inhibited tension. In the phase of rising tension, ATPase increased proportionally to tension, to 0.11 +/- 0.01 mumol X g-1 X s-1 at maximum tension. However, the ratio of ATPase to tension on the rising phase had a value only one-third of that seen with calcium-activated tension. Thus, low substrate concentrations, but not low calcium ion concentrations, influence cross-bridge kinetics under steady-state isometric conditions, possibly by an increase in the tension-time product during a cross-bridge cycle.
Regional cerebral glucose utilization measured with [14C]glucose in brief experiments
Authors: Lu D. M. , Davis D. W. , Mans A. M. , Hawkins R. A.
Published: December 2017

A method was developed to measure regional cerebral glucose utilization (rCMRGlc) in very short times using autoradiography and [2- 14C]glucose. It includes a more comprehensive mathematical treatment than prototypes and allows greater experimental flexibility. The current model takes into account the exchange of glucose between plasma and brain as a function of the plasma glucose concentration and rCMRGlc. The loss of 14C as 14CO2, although relatively small, is also calculated and the results corrected accordingly. Fully metabolizable radiolabeled glucose was chosen as the tracer, because it most closely resembles the natural substrate. Glucose is more rapidly phosphorylated than analogues such as 2-deoxyglucose and the 14C accumulated within intermediary metabolites soon account for most of the radioactivity in cerebral tissue. Furthermore, no special constant is necessary to adjust the rates as is the case when analogues are used. Excellent anatomic resolution was achieved when the technique was used to determine rCMRGlc in experiments as short as 5 min. By taking this approach, more short-lived phenomena may be studied under more diverse conditions than was previously possible.
Secretory products of macrophages and their physiological functions
Authors: Takemura R. , Werb Z.
Published: December 2017

Macrophages secrete a variety of biologically active substances into their local milieu, including proteins, lipids, nucleotide metabolites, and oxygen metabolites. To date, more than 50 substances secreted by macrophages have been reported: enzymes; enzyme inhibitors; plasma proteins such as complement components, coagulation factors, and apolipoprotein E; factors that regulate the functions of other cells such as interferon, interleukin 1, mitogens, and angiogenesis factor; and low molecular weight substances such as reactive metabolites of oxygen and derivatives of arachidonic acids. Macrophage-derived products are probably important in the local environment, and they are believed to be important in the physiological and pathological functions of macrophages in inflammation, tissue repair, lipoprotein metabolism, acute phase response, and in microbicidal, antiviral, tumoricidal, and immunoregulatory activities; however, macrophages may not be the sole source for the secretion of some of these products. The secretion of these products is intricately regulated, developmentally and environmentally.
Carotid body cell culture and selective growth of glomus cells
Authors: Fishman M. C. , Schaffner A. E.
Published: December 2017

Cells of the fetal carotid body have been obtained by enzymatic digestion and maintained in culture both as single cells and as clusters for up to 2 mo. The glomus cells in culture synthesize catecholamines and adenine nucleotides, as determined by histochemical methods, and contain characteristic dense-core vesicles. Their growth requirements are different from other cells of neural crest origin in that they do not depend on nerve growth factor (as do sympathetic neurons) or corticosteroids (as do SIF cells) for survival. Neither hypoxia nor hypercarbia affects survival or relative preponderance of glomus cells in culture. Tyrosine-free medium, which selects for cells containing tyrosine hydroxylase, eliminates most of the nonadrenergic cells, thereby providing a culture dramatically enhanced in glomus cells.
Effects of thyroid hormones on adipose tissue development in Sherman and Zucker rats
Authors: Levacher C. , Sztalryd C. , Kinebanyan M. F. , Picon L.
Published: December 2017

It has been reported that mild hyperthyroidism in the young Sherman rat induces adipose tissue hyperplasia, concomitant with cell size reduction, and that hypothyroidism induces opposite effects. The present experiments were designed to study the evolution of cellularity in retroperitoneal and epididymal adipose tissue during a long term thyroxine (T4) treatment or in T4-treated rats, after the treatment had been stopped. In both cases, hyperplasia was transient with the observation that the adipocyte number observed in 3-mo-old control rats was reached earlier in T4-treated rats. In hypothyroid rats, hypoplasia was also transient, because once the treatment was stopped, the cell number overtook that of controls. Zucker rats were also treated with T4, because hypoplasia has been observed in young obese (fa/fa) rats and these rats are reported to be hypothyroid. T4 treatment increased their adipocyte number up to the level of nonobese (Fa/fa) untreated rats, while hypertrophy, although reduced, was persistent. In Sherman and Zucker rats, adipose tissue lipoprotein lipase activity was decreased by T4 treatment in parallel with and perhaps because of adipocyte size reduction. We suggest that hyperplasia induced by thyroid hormones results from a precocious differentiation of preadipocytes and does not necessarily imply an increased preadipocytes multiplication.
Early cell alterations induced by histamine and epinephrine in rabbit lung
Authors: Gil J. , McNiff J. M.
Published: December 2017

This study was concerned with the early effects on lung cells of infusion of 18 micrograms/kg histamine to the rabbit. For comparison purposes, other rabbits received a bolus of 0.4 micrograms/kg epinephrine. After administration, the lungs were immediately fixed by vascular perfusion of osmium tetroxide. In the group that received histamine, we observed selective alterations in the type I epithelial cells of the alveolar wall, consisting of localized thickening, increased number of cytoplasmic ribosomes, and homogeneity and increased electron density with reduction of the number of plasmalemmal vesicles. These alterations were quantitated; the vesicular load (no. of vesicles/cm2) was computed separately for each cell front of epithelial and endothelial cells. Epinephrine induced only edema in epithelial and endothelial cells, resulting in high electron lucency, thickening, and irregular cell profiles. We interpret the alterations seen in histamine-treated animals as reflecting uptake of histamine by the alveolar epithelium leading to an increased level of metabolic activity. Histamine induced no interstitial edema and no immediate reduction of capillary volume. The cellular edema related to epinephrine, a substance which is not taken up or metabolized by the lung, represented a form of cell injury.
Sensitivity of cAMP-stimulated salt secretion in shark rectal gland to "loop" diuretics
Authors: Palfrey H. C. , Silva P. , Epstein F. H.
Published: December 2017

The secretory response of isolated perfused shark rectal gland was characterized with respect to its inhibition by a chemically related series of 5-sulfamoylbenzoic acid derivatives and certain phenoxyacetic acid derivatives. Maximal fluid and salt secretion was elicited with dibutyryl adenosine 3',5'-cyclic monophosphate and theophylline. The potency series established for the 5-sulfamoylbenzoic acid compounds agreed well with the relative potencies previously established for these agents as inhibitors of Na+-K+-Cl- cotransport in the avian erythrocyte. The most potent compound tested (3-benzylamino-4-phenyl-5-sulfamoylbenzoic acid) had a 50% inhibitory concentration value of approximately 5 X 10(-7) M. This compound was approximately 10-fold more effective than bumetanide and 400-fold as inhibitory as furosemide in this system. Ethacrynic acid (EA; 10(-3) M) was a poor inhibitor of rectal gland secretion and was approximately equipotent with its saturated derivative, dihydro EA. In contrast, EA-L-cysteine was fully effective at 10(-4) M, although the EA-D-cysteine adduct was ineffective. These data also qualitatively agree with results obtained in the inhibition of avian erythrocyte Na+-K+-Cl- cotransport and suggest that the rectal gland possesses a serosally oriented “NaCl cotransport” system with pharmacological and perhaps physiological similarities to the Na+-K+-Cl-cotransporter found in erythrocyte and other cell membranes.
Role of adenosine as an endogenous regulator of respiration in hamster brown adipocytes
Authors: Schimmel R. J. , McCarthy L.
Published: December 2017

The action of endogeneous adenosine on isolated hamster brown adipocytes was examined. Adenosine production from brown adipocytes was measured after labeling of the intracellular nucleotide pool with [3H]adenine. Accumulation of [3H]adenosine in the incubation medium was maximum after 5 min of incubation and was still present after 20 min. When adenosine accumulation was prevented by addition of adenosine deaminase, the stimulatory effects of isoproterenol on oxygen uptake, lipolysis, and adenosine 3',5'-cyclic monophosphate (cAMP) generation were enhanced. However, basal rates of lipolysis and oxygen consumption and levels of cAMP were not affected on addition of adenosine deaminase. A similar potentiation of isoproterenol responses was produced by the adenosine receptor antagonist, 3-isobutyl-1-methylxanthine, present at a concentration (10 microM) which did not change basal levels of respiration or lipolysis. Addition of the adenosine analogue 2-chloroadenosine antagonized isoproterenol-stimulated respiration and lipolysis and prevented potentiation of isoproterenol responses with 3-isobutyl-1-methylxanthine. To localize the site of adenosine action, activity of adenylate cyclase in membrane preparations from brown adipocytes was measured. Isoproterenol-stimulated adenylate cyclase activity was partially inhibited by 2-chloroadenosine in a GTP-dependent manner. Addition of Na+ enhanced the inhibitory effect of 2-chloroadenosine, and 3-isobutyl-1-methylxanthine blocked it. The calculated 50% effective dose for 2-chloroadenosine inhibition was between 10 and 15 nM. These data suggest that adenosine produced by brown adipocytes is an endogenous regulator of respiration in these cells acting at the level of the adenylate cyclase enzyme.
Characterization of anion transport system in trout red blood cell
Authors: Romano L. , Passow H.
Published: December 2017

Anion transport in the trout red blood cell is mediated by a membrane protein that selectively binds dihydro-4,4'-dithiocyanostilbene-2,2'-disulfonic acid (3H2DIDS) and that forms on sodium dodecyl sulfate (SDS)-polyacrylamide gel electropherograms a band with the same diffuse structure at the same location as the band 3 protein of the mammalian red blood cells. There exists a linear relationship between binding of H2DIDS to this protein and the inhibition of anion equilibrium exchange. At maximal inhibition about 8 X 10(6) molecules/cell are bound to the protein. The kinetics of anion transport in the trout red blood cell differ from those of mammalian red blood cells. In addition to a H2DIDS-sensitive component of sulfate transport there exists a considerable H2DIDS-insensitive component with a relative magnitude that decreases with increasing temperature. At 23 degrees C, it amounts to about 25%. The temperature dependence of the H2DIDS-sensitive component is about 15 kcal/mol instead of 32 as in human red blood cells. Cl- transport increases with increasing pH. Above pH 7.4, the rate of transport becomes too fast to be measurable with either inhibitor stop or filtration technique. SO2-4 transport is nearly pH independent over the pH range 6.5 to 7.8 and the net entry of SO2-4 in exchange against intracellular Cl-, as followed in the absence of CO2, is accompanied by little if any proton uptake. Net proton uptake becomes measurable only at temperatures above 40 degrees C. Possibly at lower and more physiological temperatures, the band 3 protein in the red blood cell of the trout accomplishes part of the SO2-4 movements without cotransporting protons.
Activation by N-ethylmaleimide of a latent K+-Cl- flux in human red blood cells
Authors: Lauf P. K. , Adragna N. C. , Garay R. P.
Published: December 2017

Twenty to fifty percent of the ouabain-insensitive Na+ and K+ fluxes in human red blood cells are mediated by Cl(-) -dependent coupled transport (cotransport). In this paper we report on the effect of the sulfhydryl group reagent N-ethylmaleimide (NEM) on Cl(-) -dependent ouabain-insensitive Na+ and K+ fluxes in human red blood cells. We found that NEM altered Na+ -K+ cotransport and activated a latent Cl(-) -dependent K+ transport mode normally apparently silent. This conclusion was based on the following observations. 1) At low concentrations (0.25 mM) NEM abolished the bumetanide-sensitive Na+ efflux and had no effect, even at a 10-fold higher concentration, on the bumetanide-sensitive K+ efflux. 2) At concentrations above 0.1 mM, NEM stimulated Cl(-) -dependent K+ efflux that was only partially inhibited by high concentrations of bumetanide or furosemide. In experiments using Rb+ as a K+ analogue, NEM activated Rb+ influx by stimulating the maximum velocity and lowering the apparent external cation affinity. The data suggest the presence of chemically reactive groups in human red blood cells for both Cl(-) -dependent K+ transport activated by NEM and Cl(-) -dependent coupled Na+-K+ movements.
Intracellular pH and bicarbonate activities in rabbit colon
Authors: Duffey M. E.
Published: December 2017

The short-circuited epithelium of rabbit colon is thought to actively absorb chloride ion by a mechanism in the mucosal cell membrane that exchanges chloride for bicarbonate ion. If this model is correct bicarbonate may be accumulated above electrochemical equilibrium across the mucosal cell membrane. To test this model intracellular pH was measured using a new hydrogen ion-selective liquid membrane microelectrode that is fast, highly selective, and easy to fabricate with a very small tip diameter. These measurements show that the average intracellular pH in this epithelium is 6.9 +/- 0.1. The mucosal cell membrane electrical potential difference, measured by conventional open-tipped microelectrodes, averaged -52 +/- 3 mV. Intracellular pH is above a value predicted for an equilibrium distribution of hydrogen ion across both cell membranes, implying that a mechanism exists for “uphill” extrusion of this ion from the cell. Intracellular bicarbonate activity calculated from these measurements averaged 8 +/- 1 mM. The electrochemical potential gradient for bicarbonate across the mucosal membrane averaged -28 +/- 2 mV, demonstrating that intracellular bicarbonate is concentrated above an equilibrium distribution across the mucosal membrane. Thus energy in the “downhill” electrochemical potential gradient for bicarbonate exit from the cell may drive the entrance of chloride into this epithelium and energize transepithelial chloride absorption.
Influence of calcium and cyclic nucleotides on beta-adrenergic sweat secretion in equine sweat glands
Authors: Bijman J. , Quinton P. M.
Published: December 2017

The effects of Ca2+, the cyclic nucleotides adenosine 3',5'-cyclic monophosphate (cAMP) and guanosine 3',5'-cyclic monophosphate (cGMP), and other parameters of sweat secretion from single equine sweat glands were examined in vitro. Extracellular Ca2+, the Ca2+ ionophore A23187, and the Ca2+ channel antagonist verapamil were all without effect on sweat secretion. Prolonged rinsing of the glands in Ca2+-free Ringer solution with 5 mM ethylenediaminetetraacetic acid decreased the secretion to 30% of the control sweat rate in response to the beta-adrenergic agonist isoproterenol; the sweat response was restored upon adding Ca2+ to the Ringer. cAMP but not cGMP analogues were as effective in stimulating sweat rates as isoproterenol, which elicited maximal secretory rates in vitro. cAMP stimulation was not inhibited by the beta-adrenergic antagonist propranolol. Because the equine sweat gland is predominantly stimulated via the beta-adrenergic receptor, we conclude that cAMP is a principal intracellular messenger in coupling this type of stimulus to the fluid secretion response in this tissue.
Evidence that levels of malate dehydrogenase and fumarase are increased by cAMP in rat myotubes
Authors: Lawrence J. C. , Salsgiver W. J.
Published: December 2017

We have investigated the potential role of adenosine 3',5'-cyclic monophosphate (cAMP) in controlling levels of enzymes of energy metabolism in primary cultures of rat skeletal muscle cells. Incubating myotubes with cholera toxin or forskolin (2 persistent activators of adenylate cyclase) significantly increased the levels of two enzymes of oxidative metabolism, fumarase and malate dehydrogenase. These enzymes were also increased (1.5- to 2.0-fold) by phosphodiesterase inhibitors (caffeine, theophylline, theobromine, 3-isobutyl-1-methylxanthine, papaverine, MJ 1988, Ro 20–1724, or SQ 20009) and the cAMP derivatives: 8-bromo-cAMP or dibutyryl cAMP. In contrast two enzymes of glycolytic metabolism, lactate dehydrogenase and pyruvate kinase, were not consistently affected by these agents. The results presented provide strong evidence that an increase in cAMP can lead to an increase in certain enzymes of oxidative energy metabolism.
Photoaffinity labeling of organic anion transport system in proximal tubule
Authors: Goldinger J. M. , Khalsa B. D. , Hong S. K.
Published: December 2017

The present investigation compares brush-border (BBM) and basolateral membrane (BLM) vesicles in terms of purity, function, appearance on sodium dodecyl sulfate (SDS) gels, and labeling pattern by use of N-(4-azido-2-nitrophenyl)-2-aminoethanesulfonic acid (NAP-taurine), a photoaffinity analogue of p-aminohippurate (PAH). Both BLM and BBM were vesicular by demonstration of PAH uptake into an osmotically active space and had probenecid-inhibitable uptake of PAH. Time courses for uptake were similar. 250 microM NAP-taurine resulted in a 35% inhibition of PAH uptake in BLM but it did not significantly effect PAH uptake into BBM. The latter was affected by 1 mM NAP-taurine. A comparison of Coomassie blue SDS gels of BLM and BBM showed markedly different staining patterns. A major band at approximately 52,000 daltons was more intensely stained in BLM than BBM. Major bands at approximately 40,000 and approximately 80,000 were stained more heavily in BBM than BLM. A minor protein at 26,000 in the BLM did not appear in BBM. An irreversible inhibition of PAH uptake in BLM was observed after photolysis in the presence of NAP-taurine. This was associated with the labeling of four protein bands on SDS polyacrylamide gels. In contrast no labeling was observed in BBM.
Proton-sulfate cotransport: external proton activation of sulfate influx into human red blood cells
Authors: Milanick M. A. , Gunn R. B.
Published: December 2017

Sulfate influx into human red blood cells was measured at 0 and 22 degrees C at several fixed external pH values between 3 and 10. These cells had normal internal pH and chloride concentrations so that sulfate influx was not limited by the efflux half-cycle reactions. The flux was a Michaelis-Menten function of sulfate concentration at each pH with K1/2SO4 = 4-10 mM. External protons activated influx 100-fold at a single site with a pK = 5.9 at 22 degrees C and 5.5 at 0 degrees C. This pK is similar to the value 5.99 +/- 0.3 for external proton binding to the sulfate-loaded transporter at 0 degrees C (J. Gen. Physiol. 79: 87-114, 1982). The flux was stilbene sensitive even in valinomycin-treated cells and was independent of membrane potential. This proton-activated influx appears to be proton-sulfate cotransport. At high pH there was a proton-independent flux that was membrane potential and stilbene sensitive. This proton-insensitive flux appears to be SO4(2-)/Cl- exchange or net sulfate influx. The sulfate influx over the entire pH range may be described in terms of an equation for the sum of the influxes through these two pathways on band 3.
Electron-microscopic study of the apical region of the toad bladder epithelial cell
Authors: Sasaki J. , Tilles S. , Condeelis J. , Carboni J. , Meiteles L. , Franki N. , Bolon R. , Robertson C. , Hays R. M.
Published: December 2017

Antidiuretic hormone (ADH) promotes fusion of cytoplasmic tubules with the luminal membrane and delivery of particles from the tubules to the membrane. The particles are believed to be the water-conducting elements in the membrane. We have employed several scanning (SEM) and transmission electron-microscopic (TEM) techniques to study the relationship of the cytoplasmic tubules to the luminal membrane and to the apical cytoskeleton of the toad bladder epithelial cell. This paper reports the results of freeze-crack SEM and tannic acid-fixed TEM studies, as well as studies with a resinless method of embedding. Freeze-cracked epithelial cells reveal that the tubules are anchored in a matrix of cytoskeleton and granules just below the luminal membrane, and many, if not all, retain their anchorage to the matrix after ADH-induced fusion. Tannic acid-fixed specimens show that the tubules in unstimulated cells lie horizontally. Fusion appears to involve an angulation of the tubules, and this may be the major mode of ADH-induced tubule movement. There are suggestions in the tannic acid sections of filamentous attachments of tubules to the surrounding cytoskeleton. In addition there are prominent microfilament bundles running down the microvilli and a dense concentration of filaments just below the luminal membrane. The presence of these filaments is confirmed in the resinless sections, and their possible role in ADH action is discussed.
Cerebral glucose use measured with [14C]glucose labeled in the 1, 2, or 6 position
Authors: Hawkins R. A. , Mans A. M. , Davis D. W. , Vina J. R. , Hibbard L. S.
Published: December 2017

The efficacy of [14C]glucose molecules labeled in various positions as tracers of regional cerebral glucose utilization (rCMRGlc) was examined in rats. Arteriovenous differences of different [14C]-glucose species and 14CO2 were measured across brain to determine the relative rates of 14CO2 loss. As anticipated, 14CO2 evolution decreased in the order: [U-14C]glucose greater than [2-14C]glucose greater than [1-14C]glucose greater than [6-14C]glucose. Release of 14CO2 from [6-14C]glucose was undetectable at 5 min and barely detectable at 10 min, and release from [1-14C]glucose, which includes the pentose phosphate pathway, was only slightly greater. rCMRGlc was measured with [1-14C]-,[2-14C]-, or [6-14C]glucose in 5-min experiments. The results of [1-14C]- and [6-14C]glucose were indistinguishable; no difference due to the activity of the pentose phosphate pathway was found. Both [1-14C]- and [6-14C]-glucose gave values similar to, but on the whole slightly higher than, [2-14C]glucose. It was concluded that when knowledge of total rCMRGlc is required, [6-14C]glucose is the labeled substrate of choice. When the experimental objective is measurement of energy metabolism, use of [1-14C]glucose avoids inclusion of the nonenergy-yielding pentose phosphate pathway.
Sodium-pump density of cells from dog tracheal mucosa
Authors: Widdicombe J. H. , Basbaum C. B. , Highland E.
Published: December 2017

Uptake of tritiated ouabain by cells isolated from dog tracheal epithelium showed two components: a saturable component with a Km of 5.1 X 10(-8) M and a maximal uptake of 8.3 X 10(5) molecules/cell and a nonsaturating component of uptake that was linear with concentration. Several criteria indicated that the saturable uptake component represented binding to the Na+-K+-ATPase. To estimate the average surface area per cell, a known number of cells were pelleted and weighed, and the average surface area was calculated, assuming the cells to be perfectly spherical. The validity of this assumption was confirmed by comparing the calculated surface areas of cells in isotonic and hypotonic media. From the values for maximal saturable uptake and average surface area, a pump density of approximately 2,400 sites/micron2 was calculated. Given that the apical membrane lacks Na pumps and accounts for only approximately 5% of the total surface area, this value corresponds to the pump density of the basolateral cell membrane. The pump densities of ciliated, goblet, and basal cells were compared by autoradiography. The three cell types had approximately the same density of pump sites.
Alpha 2-adrenergic receptor-mediated regulation of intestinal calcium transport
Authors: Hyun C. S. , Cragoe E. J. , Field M.
Published: December 2017

The role of alpha 2-adrenergic receptors in the regulation of intestinal Ca2+ transport was studied in vitro in rabbit proximal colon and distal ileum. Unidirectional mucosal-to-serosal and serosal-to-mucosal fluxes (Jm----s and Js----m) of 45Ca2+ were measured across isolated mucosal sheets bathed on both sides with identical Ringer solutions (1.25 mM Ca2+) and short-circuited. In the proximal colon, mucosal-to-epithelial influx (Jm----e) of Ca2+ was also determined. Proximal colonic mucosa secreted Ca2+ (in nmol X h-1 X cm-2, Jm----s = 12.1 and Js----m = 42.2). Epinephrine (10 microM) abolished Ca2+ secretion (Jm----s = 24.1 and Js----m = 24.9) but did not affect Jm----e (21.4 in controls and 23.6 after epinephrine). These effects were mediated by alpha 2-adrenergic receptors: yohimbine abolished the effect of epinephrine, but prazosin and propranolol did not; furthermore, clonidine mimicked the effects of epinephrine, whereas methoxamine and isoproterenol had no effect. Ileal mucosa did not transport Ca2+ spontaneously; epinephrine (10 microM) increased Jm----s in the ileum without affecting Js----m, thereby stimulating net Ca2+ absorption. The mechanism for these alpha 2-receptor-mediated effects was examined in proximal colon. The effect of epinephrine on Jm----s was dependent on serosal Na+ but was unaffected by ouabain or the Na+-Ca2+ exchange inhibitor, benzamil. The effect of epinephrine on Js----m was dependent on serosal Na+ and was blocked by both ouabain and benzamil. In the absence of epinephrine benzamil did not alter Jm----s or Js----m.(ABSTRACT TRUNCATED AT 250 WORDS)
Spectral characterization of ciliary beating: variations of frequency with time
Authors: Eshel D. , Grossman Y. , Priel Z.
Published: December 2017

Ciliary beating frequency in tissue culture from frog palate and isolated lung was optically examined using instrumentation that was adjusted to measure a fraction of the surface area of a single ciliary cell. Consecutive 1-s segments of the analogue signal were fast Fourier transformed (FFT) to obtain a power spectrum. At room temperature, these power spectra changed over time from 1 s to the next. Each spectrum contained several dominant frequencies of similar intensities. Cooling the preparation resulted in a single-peak spectrum that was constant over time. A mathematical model is proposed to simulate these findings. The results and the mathematical model support the hypothesis that ciliary beating frequency fluctuates over short periods of time.
Adenosine stimulates sodium transport in kidney A6 epithelia in culture
Authors: Lang M. A. , Preston A. S. , Handler J. S. , Forrest J. N.
Published: December 2017

The effects of adenosine receptor agonists and antagonists were examined in epithelia formed in culture by A6 cells, a continuous cell line derived from Xenopus laevis kidney. A6 epithelia have a high electrical resistance and a short-circuit current that is equal to net sodium flux from mucosal to serosal surface. Adenosine, 2-chloroadenosine, 5'-(N-ethyl)carboxamidoadenosine, and N6-(L-2-phenylisopropyl) adenosine produced concentration-dependent increases in short-circuit current. Stimulation of short-circuit current by 2-chloroadenosine occurred at concentrations of 0.05 microM and above, with half-maximal stimulation occurring at 0.3 microM. 5'-(N-ethyl)carboxamidoadenosine was more potent than N6-(L-2-phenylisopropyl)adenosine, the usual order of potency for activation of stimulatory adenosine receptors. Theophylline (100 microM), an adenosine receptor antagonist, reduced the short-circuit current response to adenosine and 2-chloroadenosine by 85-90%. Amiloride, an agent that inhibits both basal and adenosine 3',5'-cyclic monophosphate (cAMP)-stimulated short-circuit current in A6 epithelia, completely and reversibly inhibited short-circuit current stimulated by 2-chloroadenosine. Adenosine and 2-chloroadenosine stimulated adenylate cyclase activity in a crude membrane preparation from A6 cells. Stimulation by adenosine was blocked by adenosine deaminase. 2-Chloroadenosine increased cell cAMP accumulation in intact epithelia. The results provide evidence that adenosine and adenosine receptor agonists stimulate adenylate cyclase and active sodium transport in an epithelial cell line of renal origin.
Potassium-chloride cotransport in cultured chick heart cells
Authors: Piwnica-Worms D. , Jacob R. , Horres C. R. , Lieberman M.
Published: December 2017

The polystrand preparation of cultured chick heart cells has a unidirectional transmembrane Cl- efflux that is twice K+ efflux. However, Cl- conductance of this heart cell membrane is low [regardless of extracellular K+ (K+o)], suggesting the existence of electroneutral Cl--dependent transport mechanisms. Furosemide (10(-3) M) decreases the 36Cl tracer efflux rate constant from a control value of 0.67 to 0.33 min-1. Extracellular Na+--free solution, which depletes intracellular Na+ within 1 min, has no significant effect on 36Cl efflux. K+o-free solution plus 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS; 10(-4) M) promotes the loss of Cl- against the Cl- electrochemical gradient; Cl- loss is furosemide sensitive in a dose-dependent manner. Incubating polystrands in 133 mM K+o, normal extracellular Cl- (Cl-o) solution causes net K+ and Cl- uptake in a 1:1 stoichiometry as well as a furosemide-sensitive volume increase; 130 mM extracellular choline or Li+ cannot mimic this high-K+o-induced volume increase. Removal of Cl-o from 133 mM K+o solution prevents K+ uptake and causes a Cl- loss as well as a furosemide-sensitive volume decrease. Adjusting Cl-o concentrations in high-K+o solution plus DIDS, so that the Cl- chemical gradient equally opposes the K+ chemical gradient, prevents high-K+o-induced volume changes. These data suggest that the cardiac cell membrane contains a furosemide-sensitive K+-Cl- cotransport mechanism.
Superoxide dismutase: tissue, cellular, and subcellular distribution in adult canine heart
Authors: Spanier A. M. , Weglicki W. B. , Stiers D. L. , Misra H. P.
Published: December 2017

Cell-free extracts of canine myocardial tissue were found to contain two biochemically and electrophoretically distinct superoxide dismutases (SOD), an enzyme that provides defense against the deleterious effect of superoxide radicals (O2.-). Polyacrylamide gel (7.5%) electrophoresis revealed two distinct bands of SOD activity: a slower moving band [retardation factor (Rf) = 0.4] resembling the manganese SOD found in bacteria and mitochondria (which is not inhibited by 2.5 mM cyanide) and a faster moving band (Rf = 0.75) that is sensitive to cyanide. In contrast, extracts from isolated adult canine cardiac myocytes were found to contain only the cyanide-insensitive SOD. Extracts of whole myocardium and isolated cardiac myocytes contain 22.3 +/- 1.2 and 27.0 +/- 1.5 U cyanide-insensitive SOD/mg protein, respectively. However, the activity of cyanide-sensitive SOD in these fractions is 7.9 +/- 2.0 (tissue) and 1.5 +/- 1.4 (cells) U/mg protein. Cardiac myocyte SOD activity was particulate in nature, and the major part of the SOD activity was associated with heavy mitochondrial fractions. The biologic significance of this higher activity of SOD in the heavier mitochondrial fraction remains to be elucidated.