bluuebunny/crossref_metadata_2025 huggingface dataset:
A low-cost signal averager and data-acquisition device
Authors: Bezanilla F. , Armstrong C. M.
Published: December 2017

A low-cost data-acquisition device, which can be used as a signal averager or temporary data buffer, is described. It consists of 1) an input stage built with a multiplexer, sample-and-hold amplifier, and an analog-to-digital converter; 2) a memory section made of static shift registers with recirculate capability; 3) an output stage built with a demultiplexer and a digital-to-analog converter and, 4) a control module which provides the logic signals to operate the machine. Basic diagrams are presented and several applications are described.
Effects of temperature on some membrane characteristics of carotid body cells
Authors: Baron M. , Eyzaguirre C.
Published: December 2017

Excised cat carotid bodies and nerves were placed in a Lucite chamber through which saline flowed at different temperatures. Action potentials were recorded with a suction electrode. Glomus cells were impaled with microelectrodes and identified by ejecting Procion navy blue from the micropipette. Membrane potentials (MP) averaged 20 mV and input resistances (R0), 40 Momega. Cooling induced depolarization and decreased R0. This effect had a reversal potential of about -6 mV. [K+]0 did not affect MP, R0, or cooling effects. Lack of sodium increased MP, R0, and depressed cooling effects. Low chloride had opposite effects. Ca2+ and Mg2+ influenced MP and R0 but had little effects on temperature actions. Ouabain decreased MP and R0. Results indicate that glomus cells are exquisitely sensitive to temperature changes; their MP and R0 increase with high temperatures and decrease when temperature is lowered. These effects are mostly dependent on the external concentration of Cl- and Na+. Glomus cells may be the site of the intense metabolism of this tissue and thus contribute to the generation of chemosensory impulses.
Excitation-contraction coupling in multiunit tracheal smooth muscle during metabolic depletion: induction of rhythmicity
Authors: Bose R. , Bose D.
Published: December 2017

Multiunit canine tracheal smooth muscle responded to carbachol with graded depolarization and tonic contraction. The same concentration of carbachol, after metabolic depletion by substrate removal, produced rhythmic contractions and action potentials. Similar mechanical effects were also observed with acetylcholine or histamine. These effects were reversed by reintroducing glucose or beta-hydroxybutyrate, but not by 3-O-methylglucose, which is not metabolized; hence, the structural requirements for glucose, per se, or any osmotic effect were ruled out. Sensitivity to extracellular Ca2+ was increased. A Ca2+-influx blocker, D-600, in low concentration (2 X 10(-8) M) abolished the rhythmic contractions without affecting the tonic contraction. Progressive metabolic depletion in presence of carbachol led to fluctuations in membrane potential with a crest of depolarization and appearance of action potentials, each of which resulted in a small contraction. Many of the small contractions partially fused to form the major rhythmic contractions which appeared at a frequency of one per minute. Rhythmicity could not be produced by increasing extracellular K+ concentration (20-120 mM) in presence of atropine (13(-7) M), but instead a tonic contraction occurred. These results suggest changes in excitation-contraction coupling mechanism with agonists like acetylcholine, carbachol, or histamine during substrate deprivation.
Mechanical properties of tracheal smooth muscle: effects of temperature
Authors: Stephens N. L. , Cardinal R. , Simmons B.
Published: December 2017

The effect of temperature on the isometric tetanic myogram was studied in isolated canine tracheal smooth muscle (TSM). At 37 degrees C and 27 degrees C no significant change occurred in maximum tetanic tension (PO). At 17 degrees C a significant reduction was seen Values of Q10 for contraction time (tPO) were almost halved, whereas those for rate of tension development (dP/dt) were almost doubled. The effect of the same temperatures on the force-velocity (F-v) relationships was also studied. All three F-v curves were described by the Hill equation, (P + a) (v + b) = (PO + a)b. Vmax and b decreased with decreased temperature, with Q10's demonstrating they were dependent on active processes. Finally, the decreased dP/dt of the myogram at lower temperatures was felt to be the probable result of decreased contractile element velocity because no decrease in series elastic component stiffness was demonstrable, there being instead an increase in stiffness at lower temperatures.
Bioelectrical and contractile behavior of rabbit mesotubarium superius: estrogen withdrawal
Authors: Conrad J. T.
Published: December 2017

The mesotubarium superius of the female rabbit is a thin, plane sheet of smooth muscle responding to the in vivo endocrine states of ovulation, estrous, castration, pregnancy, and replacement estrogen therapy of the castrate in a manner similar to that of the uterus. Both isometric tension and microelectrode recorded transmembrane potentials were measured in vitro to make these comparisons. In addition, confirmation of the “estrogen-withdrawal” theory of activation was obtained when it was noted that about 12 h after an abrupt withdrawal of estrogen there is an increase in contractile activity coinciding with an increase in subthreshold activity and a slight (about 5 mV) decline in the average resting potential compared to the control estrous level. It is concluded that the mesotubarium superius is an excellent tissue for reproductive smooth muscle studies, where a geometrically simple model is useful.
Placental amino acid uptake. IV. Transport microvillous membrane vesicles
Authors: Ruzycki S. M. , Kelley L. K. , Smith C. H.
Published: December 2017

The recent characterization of a vesicular microvillous plasma membrane preparation from human placenta has made possible study of the characteristics of amino acid transport by the isolated membrane. Variation of medium osmolality demonstrated that alpha-aminoisobutyric acid (AIB) is taken up virtually entirely into a vesicular space. Vesicular volume was determined from equilibrium uptake of three dissimilar nonamino acid substrates, all of which gave a common value. Temperature dependence, saturability, specificity, and similarity of kinetic constants all indicated that AIB uptake by the vesicles resembles that by whole villous tissue. AIB uptake was stimulated specifically by an inward sodium gradient and unaffected by an outward potassium gradient. The persistence of stimulation up to 2 h after the gradient was established could be correlated with the slow equilibration of sodium across the membrane. AIB exodus could be studied with minimal problems of substrate recapture by using preloaded membrane vesicles bound tightly to cellulose ester filters to facilitate transfer to fresh medium. The exodus process was saturable with a Km considerably greater than that for entry.
Controlled bending of high-resistance glass microelectrodes
Authors: Hudspeth A. J. , Corey D. P.
Published: December 2017

The short working distance of compound microscope lenses in many cases prevents microelectrode penetrations of cells perpendicular to the cell surface. Bending electrodes very near their tips removes this constraint. A method is described for bending glass microelectrodes with a hot filament while their tips are immersed in a water drop. Immersion protects the fine electrode tips from the heat and provides control over the angle through which the electrodes are bent.
Magnesium net fluxes and distribution in rabbit myocardium in irreversible contracture
Authors: Paradise N. F. , Beeler G. W. , Visscher M. B.
Published: December 2017

Distribution of magnesium (Mg) in heart muscle was studied by measuring fluxes of Mg and transmembrane potentials as a function of perfusate [Mg2+] after a massive increase in permeability of the sarcolemma was induced in the Langendorff prepared heart from the Nembutal-anesthetized rabbit. After onset of 0 mM [Ca2+] perfusion which produced excitation-contraction (E-C) uncoupling and mechanical arrest, action potentials recorded from subepicardial cells showed an increase in duration and decrease in amplitude, which progressed until no transmembrane potentials could be observed. Restoration of physiological salt solution perfusion after 15 min of [Ca2+]-free perfusion caused an irreversible contracture that was associated with 1) efflux of potassium (K) and myoglobin, 2) perfusate [Mg2+]-dependent flux of Mg, and 3) transmembrane potentials of 0 mV. The magnitude of net efflux of K and myoglobin during contracture was unaffected by perfusate [Mg2+]. During the first 2 min of contracture, net efflux of Mg (mumoles per gram wet muscle +/- SE) was 1.37 +/- 0.09 and 0.48 +/- 0.19 during 0 mM and 2.5 mM [Mg2+] perfusion, respectively; but a net influx of 0.56 +/- 0.23 occurred during 5 mM [Mg2+] perfusion. Total sarcoplasmic [Mg] may correspond to perfusate [Mg2+] of 3.6 mM, which was found by interpolation to prevent any net flux of Mg during contracture. 3.6 mM may, therefore, represent the upper limit of the intracellular free-ionized Mg concentration in rabbit heart.
Adrenergic influences on rabbit oviduct: effect of muscle size and ovarian hormones
Authors: Meiri H. , Meiri U. , Kennedy D. R. , Marshall J. M.
Published: December 2017

This study attempts to distinguish between a direct action of ovarian steroids on adrenergic neurons in the oviduct and an indirect effect mediated by changes in muscle size. Mature rabbits were treated as follows: group 1, ovariectomized and a polyethylene catheter (1 mm OD) inserted into isthmus of one oviduct (CT) with contralateral oviduct as control (C); group 2, normal, estrous animals with one intubated isthmus (ET) and contralateral control (E). Fourteen days postoperatively, oviducts were removed and muscle-wall thickness, norepinephrine (NE) content, and response to nerve stimulation were measured. Although castration atrophy was prevented in CT, NE content of CT was significantly less than C, 0.032 +/- 0.07 versus 1.09 +/- 0.10 nmol. NE content of ET was also significantly less than E (1.32 +/- 0.03 versus 1.81 +/- 2.0) despite a greater wall thickness of ET. It was concluded that: a) withdrawal of ovarian hormones reduces NE contents by a direct action on nerves; b) moderate stretch, per se, increases muscle size and reduces NE content; c) nerve stimulation induces muscle contraction despite large reduction in transmitter content.
Herpes virus infection alters electrical parameters of heart cell aggregates
Authors: Shrier A. , Nahmias A. J. , DeHaan R. L.
Published: December 2017

The spontaneous beat of embryonic heart cell aggregates in culture grew gradually weaker and stopped about 18 h after inoculation with herpes simplex virus 1 (HSV-1). Before electrical activity ceased, maximal action potential upstroke velocity fell to about 20% of control values, whereas overshoot and plateau duration declined about 50%. Maximal diastolic potential was reduced by only about 10%. Eighteen hours after viral inoculation, specific membrane resistance and intercellular coupling were measured in quiescent aggregates with injected current pulses passed between two widely spaced intracellular electrodes. These parameters were unaltered as compared with control aggregates. However, pairs of infected aggregates brought into contact required 8 h to synchronize their beats; mock-inoculated aggregates coupled in less than 1 h. It is concluded that the cell surface alterations caused by HSV-1 infection specifically reduce both fast and slow inward currents and interfere with the formation of new nexal junctions.
Calcium fluxes and contractility in isolated guinea pig atrium: effect of A23187
Authors: Holland D. R. , Armstrong W. M. , Steinberg M. I.
Published: December 2017

The Ca2+ ionophore A23187 (10(-6) to 3 X 10(-5) M) increased the force of contraction is isolated guinea pig atria. In individual twitches, peak tension, maximum rate of tension development, time to peak tension, and total twitch duration were all increased by A23187. Tripelennamine, indomethacin, and atropine did not significantly alter the inotropic effect of A23187. Serotonin produced changes in individual twitches that differed qualitatively and quantitatively from those of A23187. Therefore, the inotropic action of A23187 is probably not mediated by release of endogenous histamine, prostaglandins, acetylcholine, or serotonin. 45Ca influx and efflux were increased by A23187. The enhanced 45Ca efflux exceeded that which would be predicted if the ionophore acted only to increase the passive Ca2+ permeability of the myocardial cell membrane. These results suggest that A23187 facilitates the entry of extracellular Ca2+ into the myocardial cell and the release of intracellular Ca2+ stores into the myoplasm. The resultant increase in intracellular Ca2+ activity could account for the positive inotropic action of A23187.
Energy coupling to sodium transport in frog skeletal muscle
Authors: Connett R. J. , Hays E. T.
Published: December 2017

In addition to a strophanthidin-sensitive (SS) sodium efflux, a large component of the sodium efflux in freshly isolated frog skeletal muscle is sodium-activated and strophanthidin-insensitive (SASI). The amount of metabolic energy associated with sodium movement by each of these components was measured and the coupling between sodium movement and adenosine 5'-triphosphate (ATP) hydrolysis in muscle was calculated. Energy production was blocked by iodoacetate and cyanide. Energy turnover was estimated from the change in creatine phosphate (CrP) and ATP contents and expressed as potential energy (PE = CrP + 2ATP). After metabolic poisoning a linear fall of PE occurred (6.3 mumol/g.h). Metabolic poisoning had no effect on the magnitude of the SS or SASI components of sodium efflux. In 2 h the sodium moved, and PE change due to the SS component was 4.35 and 1.66 mumol/g.h, respectively, which gave a coupling factor of 2.6. The amount of sodium moved by the SASI component was similar to that moved by the SS component in 2 h whereas no energy change was observed. It was, therefore, concluded that sodium movement by the SASI component requires no energy input.
Reactive forms of oxygen and chemiluminescence in phagocytizing rabbit alveolar macrophages
Authors: Miles P. R. , Castranova V. , Lee P.
Published: December 2017

Chemiluminescence (CL), superoxide anion (O2-) production, and particle uptake were measured to determine the role of antibacterial substances in the chemiluminescent response associated with phagocytosis in rabbit alveolar macrophages (AM). Exposure of AM to zymosan particles induced both CL and the production of extracellular O2-. CL is inhibited by superoxide dismutase, an enzyme which catalyzes the conversion of O2- to hydrogen peroxide (H2O2), by catalase, an enzyme which destroys H2O2, and by the hydroxyl radical (.OH) scavengers, benzoate and ethanol. Superoxide dismutase and catalase probably exert their effects in the extracellular fluid. CL can also be produced by the addition of NaO2 or H2O2 to zymosan in a noncellular system. The chemiluminescent response occurs before particle uptake is complete, which also indicates that CL occurs in the extracellular fluid. These results suggest that CL induced by zymosan in AM is due to the extracellular reaction between various reactive forms of oxygen and zymosan.
Further similarities of endogenous pyrogen and leukocytic endogenous mediator
Authors: Kampschmidt R. F. , Pulliam L. A. , Merriman C. R.
Published: December 2017

The release of endogenous pyrogen (EP) from rabbit peritoneal granulocytes was measured with a three-point log dose-response curve. Release of EP was inhibited when the cells were incubated in media containing potassium or calcium. Measurements of leukocytic endogenous mediator (LEM) activity, i.e., lowering of plasma iron and zinc and increases in blood neutrophils, were made on the same supernatant media. When EP release was inhibited there was a similar inhibition of LEM activity. These results indicate a similarity between the release of pyrogenic and LEM activities. Together with previous purification studies, the results suggest that EP and LEM are similar and may be identical factors.
Quantitative ultrastructural analysis in cardiac membrane physiology
Authors: Page E.
Published: December 2017

Quantitative measurements on electron micrographs of heart muscle can yield information useful for cellular physiologists and at present not obtainable in other ways. These methods are subject to preparative artifact, sampling problems, and problems inherent in the mathematical description of ultrastructure. Nevertheless they provide the best available data for membrane areas of the plasmalemma and its components, as well as for membrane areas of the sarcoplasmic reticulum and mitochondria. Morphometric methods can be used to study growth of membranes. Changes in the volumes of intracellular membrane-limited subcompartments can also be measured. Quantitative analysis of freeze-fractured membrane replicas can be carried out either by a statistical approach or by optical diffraction. In this way, physiological perturbations or developmental events leading to changes in membrane permeability can be studied for correlated changes in membrane structure.
Ionic milieu and control of K permeability in the parotid gland
Authors: Putney J. W.
Published: December 2017

The roles of Mg, K, Na, and Cl ions in the biphasic K-permeability response (86Rb release) to receptor activation in the parotid gland were investigated. Both the transient and sustained phases of K release (86Rb release) were unaffected by alterations in Mg concentration. Elevating extracellular K enhanced 86Rb release presumably by decreasing the negativity of the membrane potential. Elevating extracellular K had no effect on the transient or sustained phases of 86Rb release due to carbachol and 1.0 mM Ca. Measurements of responses to carbachol with various substitutes for NaCl suggested that extracellular Cl and ionic strength are important for the K-permeability response. Experiments with LiCl suggested the operation of a Na-Ca exchange similar to that seen earlier in experiments with amylase release. When tetraethylammonium Cl was substituted for NaCl, the cation appeared to block K fluxes directly. Choline Cl, in partial substitution for NaCl, potentiated the sustained phase of response to substance P and, in complete substitution, produced both Ca-dependent and Ca-independent increases in 86Rb release. It was concluded that the 86Rb-release response in relatively (but not completely) insensitive to alteration in the ionic milieu. It was also concluded that Na regulates K permeability in part directly and in part by means of a Na-Ca exchange.
Differences in ammonia and adenylate metabolism in contracting fast and slow muscle
Authors: Meyer R. A. , Terjung R. L.
Published: December 2017

The time course of AMP deamination and IMP reamination was studied during in situ stimulation and subsequent recovery in fast-twitch (gastrocnemius) and slow-twitch (soleus) muscles of pentobarbital-anesthetized rats. Muscles were stimulated tetanically at rates that initially produced comparable decreases (40%) in tension development. In fast muscle, progressive decreases in total adenine nucleotide (TAN) contents of up to 50% were balanced by equivalent increases in IMP contents. Ammonia concentration initially increased in a 1:1 stoichiometry with changes in IMP and TAN. During recovery following stimulation, IMP removal matched, but NH3 removal exceeded the rate of TAN resynthesis. In contrast, TAN content in slow muscle was only slightly decreased (10%) during stimulation and there were no increases in IMP or NH3. Stimulation of the soleus following ligation of the blood supply did not increase TAN depletion. In both fast and slow muscle, changes in glutamate, aspartate, and alanine could be accounted for by transamination. These results illustrate a fundamental difference in adenylate metabolism during intense muscle stimulation between fast- and slow-twitch mammalian muscle.
Chloride distribution and exchange in rat ventricle
Authors: Polimeni P. I. , Page E.
Published: December 2017

The cellular Cl content and concentration ([Cl]cell) and the cellular uptake of 36Cl have been measured in the rat left ventricle in vivo. The in vitro efflux of 36Cl from perfused contracting ventricles preequilibrated with 36Cl in vivo was also determined at 22, 30, and 38 degrees C. [Cl]cell was 8.2 +/- 0.5 mmol/kg cell water, corresponding to a calculated equilibrium potential of Cl of -70 to -80 mV. This figure for [Cl]cell is significantly lower than previous estimates in the literature, which were subject to an analytical error leading to overestimation of muscle Cl content obtained coulometrically. At 38 degrees C, Cl exchange under quasi-steady-state conditions was 31.2 mumol . (g dry ventricle . min)-1 or 42.5 pmol . (cm2 plasma membrane . s).-1 Apparent activation energy of the flux was 10.4 kcal/mol. At 22 degrees C, no dependence of the exchange on contraction frequency was detectable over a range of 80-160 contractions/min. The Cl exchange flux is among the fastest, if not the fastest. known for myocardial ion transport.
alpha-Methylglucoside satisfies only Na+-dependent transport system of intestinal epithelium
Authors: Kimmich G. A. , Randles J.
Published: December 2017

The unidirectional influx of alpha-methylglucoside (alpha-MG) by isolated chicken intestinal epithelial cells is 98% inhibited by phlorizin. The remaining 2% of the total influx occurs in the absence of Na+, is not sensitive to phloretin, and is equal to the diffusional entry rate for 2-deoxyglucose. The glucoside is much more strongly accumulated (75-fold) than 3-O-methylglucose (3-OMG) (10-fold). Inhibitors of the serosal sugar carrier (phloretin, cytochalasin B, theophylline, and flavanoids) do not enhance alpha-MG accumulation. It is concluded that the glycoside is not a substrate for the intestinal serosal transport system. Steady-state gradients of the sugar can be represented accurately by a concentrative, phlorizin-sensitive system that is opposed by a diffusional efflux process.
Contraction of arterial smooth muscle induced by magnesium ions
Authors: Ohhashi T. , Azuma T.
Published: December 2017

Dose-response relationships for Mg2+ in a Mg2+-free bathing solution were obtained with isolated canine femoral arteries. At concentrations less than 5 mM, relaxant responses appeared in the preparations. In a concentration range of 10-20 mM, however, every one of the preparations invariably showed dose-dependent contractions. The relaxant and contractile responses were induced by the action of Mg2+ per se but not by the hyperosmolarity associated with the addition of Mg2+. Neither response was affected by the drugs that inhibited the actions of catecholamine, acetylcholine, serotonin, or histamine. The contractile response was kept unchanged in Ca2+-free Locke's solution and was almost independent of environmental calcium concentrations ranging from 2.2 to 8.2 mM. The contractile response was not affected under the influence of Ca antagonists, verapamil, or Mn2+. The presence of Ba2+ or Sr2+ enhanced the relaxant response and dose dependently suppressed the contractile response. The contractile response was still evoked in smooth muscles that were in the state of K+ contracture. These results suggest the possibility that the contractile action of Mg2+ was mediated by a release of intracellular Ca2+ or exerted directly on the contractile machinery of arterial smooth muscle.
Intracellular oxygen supply during hypoxia
Authors: Jones D. P. , Kennedy F. G.
Published: December 2017

The intracellular supply of O2 to mitochondria was studied in single-cell suspensions of rat hepatocytes by measuring the O2 dependence of oxidation of cytochromes. Values were obtained by adding standardized O2-containing solutions to anaerobic cell suspensions and observing absorbance changes at wavelength pairs specific for cytochromes a + a3, c + c1, and b561 + b566. Half-maximal oxidation, calculated relative to aerobic cells, occurred at 3.5, 6.2, and 4.9 microM O2, respectively. These values are similar to those for changes in cellular ATP/ADP, lactate/pyruvate, and NADH/NAD+ but are much higher than corresponding values for isolated mitochondria. Analysis of these data indicate that the diffusion coefficient in the region of mitochondria in situ is considerably smaller than the extracellular diffusion coefficient and suggests that a significant O2 gradient in the vicinity of mitochondria occurs under hypoxic conditions. These results suggest that the rate of O2 diffusion may be a critical factor for intracellular O2 supply to mitochondria during hypoxia.
Blood oxygen consumption and erythrocyte types in embryonic and postnatal chicken
Authors: Grima M. , Girard H. , Dejours P.
Published: December 2017

In an extension of previous work showing that oxygen consumption (MO2) of chick blood cells declines from 5 to 14-15 days incubation age, declines more steeply to hatching, rises abruptly up to 4-6 posthatch days, and declines thereafter, we investigated the succession of embryonic and posthatch erythrocytic types by performing differential erythrocyte (RBC) counts. MO2 of RBC groups separated by density gradients and fractionation into younger and older RBC populations were measured in 1- to 22-day-old posthatch chicks. The embryonic MO2 decline can be attributed to the appearance of erythrocytic types with successively lower metabolism. The MO2 slope change at 14-15 days incubation coincides with the predominance of definitive erythrocytes with oval nuclei over other cell types. The posthatch MO2 rise is due to a temporary invasion of immature RBC, especially postnatal polychromatic erythroblasts. MO2 then decreases as these cells mature.
Bumetanide inhibits (Na + K + 2Cl) co-transport at a chloride site
Authors: Haas M. , McManus T. J.
Published: December 2017

Chloride-dependent cation transport systems in a number of cells and tissues are inhibited by 5-sulfamoylbenzoic acid loop diuretics, such as furosemide and bumetanide. Interactions between chloride and bumetanide have been examined in the catecholamine-activated (Na + K + 2Cl) co-transport pathway of the duck red blood cell. Levels of chloride were varied while maintaining a constant ratio of internal to external chloride across the cell membrane. Increasing external chloride from 20 to 100 mM shifted the dose-response curve for the effect of bumetanide on co-transport toward higher concentrations of the drug. The bumetanide concentration producing half-maximal inhibition (IC50) was increased from approximately 6 X 10(-8) to approximately 2 X 10(-7) M. When cells were incubated in the presence of a constant, submaximal inhibitory dose of bumetanide (10(-8) M), increasing external chloride (in increments of 20 mM) from 20 to 140 mM progressively decreased the level of inhibition of the co-transport system. Kinetic analysis of the data demonstrates that bumetanide and chloride compete for a common site.
Ca2+, myosin phosphorylation, and relaxation of arterial smooth muscle
Authors: Gerthoffer W. T. , Murphy R. A.
Published: December 2017

Relaxation of tissues prepared from the swine carotid media following agonist (110 mM K+) washout was analyzed as a dual-exponential decay. The time course of the initial rapid phase (about 2 min) corresponded to myosin dephosphorylation and to the decay of the capacity to shorten isotonically. Because myosin was dephosphorylated to basal levels within 2 min, we hypothesize that the later, slow phase of relaxation (lasting up to 45 min) was due to a slow inactivation of nonphosphorylated cross bridges. Removing extracellular Ca2+ (0 mM CaCl2, 0.1 mM ethyleneglycol-bis(beta-aminoethylether)-N,N'-tetraacetic acid) greatly enhanced the rate of the slow phase of relaxation, and raising extracellular CaCl2 to 5 mM slowed relaxation significantly. A slow rate of Ca2+ removal to a final concentration that maintains resting tone appears to produce the slow phase of relaxation. These results support hypotheses based on other studies of contracting muscles. There appear to be two populations of cross bridges interacting with the thin filament: 1) phosphorylated and capable of rapid cycling, and 2) dephosphorylated cross bridges that can maintain stress. The latter reflect an unidentified regulatory mechanism, which appears to have a high sensitivity for Ca2+.
Muscle fiber regeneration in nerve-intact and free skeletal muscle autografts in cats
Authors: Maxwell L. C.
Published: December 2017

Extensor digitorum longus (EDL) muscles of adult cats were transplanted as free autografts (FRA) with the nerve severed or as nerve-intact autografts (NIA) with the nerve retained. Histochemical and contractile properties of NIA and FRA were analyzed at selected times from 1 to 14 wk after surgery. Regeneration was qualitatively similar in NIA and FRA. Regenerating fibers were observed in both NIA and FRA within 2 wk. After 14 wk there were fewer type I fibers in both NIA and FRA than in control EDL muscles. Capillarity was greater in NIA than FRA, but both types of autografts had significantly reduced capillarity relative to control muscles. Mean fiber area, muscle mass, and absolute tension development were greater in NIA than FRA but did not reach control muscle values. Muscle mass, mean fiber area, and contractile properties, but not the proportion of type I fibers, develop toward control values more quickly in autografts with the nerve left intact.
A simple analysis of the "phosphocreatine shuttle"
Authors: Meyer R. A. , Sweeney H. L. , Kushmerick M. J.
Published: December 2017

The diffusive mobility of solutes chemically connected by reversible reactions in cells is analyzed as a problem of facilitated diffusion. By this term we mean that the diffusive flux of any substance, X, which is in one metabolic pathway, is effectively increased when it participates in a second and equilibrium reaction with another substance Y because the total flux of X in the pathway is the sum of the fluxes of X and Y. This notion is generalized and is seen to include the familiar enhanced intracellular diffusion of oxygen by oxymyoglobin. In this framework the function of creatine kinase (CK) is seen to have two aspects: 1) phosphocreatine (PCr) via the CK reaction buffers the cellular ATP and ADP concentrations and 2) transport of high-energy phosphates is predominantly in the chemical form of PCr. This predominance of PCr is a consequence of the maintained ATP, ADP, and total creatine levels and of the apparent equilibrium constant of the reaction. Thus experimental results demonstrating the transport aspects of the CK reaction emphasize only one feature of a more general notion of facilitated diffusion by near-equilibrium metabolic reactions and do not per se establish the existence of any physical or functional compartmentation of ATP, ADP, PCr, or creatine. PCr can be a large source for increasing inorganic phosphate levels during contractile activity, possibly as a metabolic regulator. Neither the transport nor buffer aspects can be quantitatively important in cells with small distances between ATP-utilizing and ATP-generating sites, such as is the case with cardiac myofibrils and mitochondria.
Postnatal development of the bronchiolar Clara cell in rats
Authors: Massaro G. D. , Davis L. , Massaro D.
Published: December 2017

We studied aspects of perinatal rat bronchiolar Clara cell development. We found that the volume density (Vv) of glycogen areas decreased 10-fold between postnatal days 1 and 2 and that the Vv of secretory granules, rough endoplasmic reticulum (RER), and mitochondria increased markedly during the 1st postnatal wk. Compared with pups of dams allowed food ad libitum during gestation, pups from dams underfed during gestation had a higher Vv of glycogen areas for the first 2 postnatal days but a lower Vv of secretory granules and RER for the time studied (to age 7 days). Glucagon injected, in utero, into fetal rats at 21.5 days of gestation resulted in a 40% decrease in the Vv of the glycogen areas within 4 h. We conclude that 1) bronchiolar Clara cells are immature at birth and undergo marked early postnatal maturation, 2) prenatal events (maternal undernutrition) affect postnatal maturation of the Clara cells, and 3) exogenous glucagon leads to glycogen depletion in Clara cells of fetal rats of 21.5 days gestation.
Involvement of calcium and cytoskeleton in gallbladder epithelial cell volume regulation
Authors: Foskett J. K. , Spring K. R.
Published: December 2017

The importance of calcium and cellular cytoskeletal elements in the activation or control of volume regulation by epithelial cells was explored in Necturus gallbladder. Gallbladder cells have been previously shown to rapidly readjust their volumes to control size after osmotic perturbation of the mucosal bathing solution. Removal of calcium from the perfusates caused dramatic morphological changes that prevented assessment of the role of extracellular calcium in volume regulation. The regulatory volume increase (RVI) that follows shrinkage of the cell due to perfusion of a hypertonic mannitol solution is insensitive to agents that interfere with cell calcium- or calmodulin-mediated events (quinidine, trifluoperazine) and is not blocked by agents that cause changes in the cytoskeleton (colchicine, cytochalasin B). Osmotically induced cell swelling is followed by regulatory volume decrease (RVD), which is inhibited by agents that interfere with calcium-dependent processes (quinidine, trifluoperazine) and by the microfilament inhibitor, cytochalasin B. These results indicate that RVD depends on calcium, calmodulin, and an intact microfilament network, whereas RVI is independent of these factors.
Synthesis and binding of lactose-specific lectin by isolated lung cells
Authors: Whitney P. , Maxwell S. , Ryan U. , Massaro D.
Published: December 2017

Neonatal maturation of alveolar structure in rat lung temporally coincides with the peak activity of a beta-galactoside-specific endogenous lung lectin of the type that has been linked to developmental processes in other tissues. To learn more about the lung lectin we examined four cell types for their ability to synthesize and bind the lectin. Cultured lung fibroblasts and pulmonary artery endothelial cells synthesized and bound the lectin, alveolar macrophages bound but did not synthesize it, and alveolar epithelial type II cells neither bound nor synthesized it. Pulmonary macrophages made a different lactose-binding protein that did not agglutinate or bind trypsin-treated red blood cells, a property of lung lectin that is the means of its assay.
Cell volume and metabolic dependence of NEM-activated K+-Cl- flux in human red blood cells
Authors: Lauf P. K. , Perkins C. M. , Adragna N. C.
Published: December 2017

The effects of incubation in anisosmotic media and of metabolic depletion on ouabain-resistant (OR) Cl--dependent K+ influxes stimulated by N-ethylmaleimide (NEM) were studied in human red blood cells using Rb+ as K+ analogue. The NEM-stimulated but not the basal Rb+-Cl- influx measured in phosphate-buffered anisosmotic media was found to be cell volume dependent. When cellular ATP, [ATP]c, was lowered to less than 0.10 of its initial level by exposure to nonmetabolizable 2-deoxy-D-glucose, the NEM-stimulated but not the basal Cl--dependent Rb+ influxes were abolished. Metabolically depleted red blood cells subsequently repleted by incubation in glucose plus inosine regained the NEM-inducible Rb+ (K+) transport activity. The difference in the time course of ATP breakdown and Rb+ influx inhibition suggests that energization of the NEM-stimulated Rb+ flux by metabolism may involve factors additional to ATP.
Control of gap junction formation in canine trachea by arachidonic acid metabolites
Authors: Agrawal R. , Daniel E. E.
Published: December 2017

This study examined whether the synthesis of the metabolites of arachidonic acid (AA) was involved in gap junction formation by 4-aminopyridine (4-AP) treatment in vitro in canine trachealis. Studies were made of the effects on gap junction formation of putative inhibitors of the cyclooxygenase and of both this and the lipoxygenase pathway of AA metabolism and the direct effects of prostaglandins (PG) E2 and I2. The number of gap junctions of similar size was increased after brief exposure to 4-AP. After indomethacin (IDM), 4-AP treatment decreased the number of gap junctions but did not affect their size. Pretreatment with 5,8,11,14-eicosatetraynoic acid or nordihydroguiaretic acid, putative inhibitors of cyclooxygenase and lipoxygenase enzymes, inhibited both the 4-AP-induced increase and decrease in the number of gap junctions. FPL 55712, a putative antagonist of leukotriene C4, did not alter either the number or the size of gap junctions when added alone or in combination with IDM. AA alone increased the number of gap junctions, but after IDM, AA decreased the number of gap junctions compared with the controls. Incubation of trachealis strips in vitro for 30 min with PGE2 increased the number of gap junctions by about threefold along with an increase in the size of the gap junctions. Similar incubation with PGI2, however, increased the number of gap junctions by approximately 60% without any change in the size. In the course of some control experiments, an interaction between carbachol and alcohol was observed such that alcohol caused an IDM-sensitive relaxation of carbachol-induced contractions, which was not observed when serotonin was the contractile agent. These results strongly suggest that PGE2 and PGI2 increase the formation of gap junctions in canine trachealis and that these prostanoids are released by 4-AP treatment. Leukotrienes may also be inhibitory in the formation of gap junctions, but FPL 55712 did not affect either the increase or the decrease in gap junctions after 4-AP.
Force-velocity curves for smooth muscle: analysis of internal factors reducing velocity
Authors: Seow C. Y. , Stephens N. L.
Published: December 2017

In tracheal smooth muscle, we obtained quantitatively different force-velocity (F-V) curves at early (2 s) and late (8 s) stages of an isometric tetanus whose contraction time was 12 s. These were essentially two samples from a continuum of F-V curves operating between 0 and 12 s. The cross-bridge cycling velocity at 8 s was slower and less sensitive to external load change compared with that at 2 s. This is possibly due to the presence of two types of cross bridges with different F-V characteristics; at 2 s most of the bridges resemble a cycling type, whereas at 8 s there is a population of what Dillon et al. have called slowly cycling or latch type interactions. Another possibility is that, due to some intrinsic factors, the whole population of cross bridges gradually change their F-V characteristics. Functions a(t) and b(t) were obtained by applying load clamps at 1-s intervals throughout a tetanus (a and b are asymptote values derived from the F-V hyperbolic curves): a increased with time, b remained constant. Analysis suggested that a/b was a valid index of internal factors that affect shortening velocity of unloaded muscle, and it is progressively increased in value during contraction.
Adenylate cyclase regulation in intact cultured myocardial cells
Authors: Marsh J. D. , Roberts D. J.
Published: December 2017

To examine the coupling of cardiac cell-surface beta-adrenergic receptors to adenylate cyclase activation and contractile response, we studied this receptor-effector response system in monolayers of spontaneously contracting chick embryo ventricular cells under physiological conditions. The hydrophilic ligand 3H-CGP12177 identified uniformly high-agonist affinity beta-adrenergic receptors (isoproterenol KD = 15 +/- 9 nM). Isoproterenol-stimulated cyclic AMP (cAMP) accumulation with 50% effective concentration at (EC50) = 12.1 nM and augmented contractile response with EC50 = 6 nM under identical conditions. One micromolar isoproterenol induced receptor loss from the cell surface with t1/2 = 13.2 min; under identical conditions cAMP content declined with t1/2 = 13.5 min and contractile response with t1/2 = 20.7 min. After agonist removal cAMP response recovered with t1/2 = 15.7 min and receptors with t1/2 = 24.7 min. Sixty minutes after agonist removal there was recovery of 52% of maximal cAMP responsiveness and 82% of the initial number of receptors; receptor occupancy was associated with 78% of initial contractile response. Agonist affinity for cell-surface receptors was changed only modestly by agonist exposure. We conclude that for this system there is relatively close coupling between high-affinity receptors, adenylate cyclase stimulation, and contractile response.
Cation dependence of posttetanic potentiation of neuromuscular transmission
Authors: Misler S. , Falke L. , Martin S.
Published: December 2017

We have investigated the possibility that much of posttetanic potentiation (PTP) of quantal release of neurotransmitter at the frog neuromuscular junction may be due to posttetanic accumulation of [Ca2+]i, via a plasmalemmal Cao2+-Nai+ exchanger that is powered by an increase in Nai+ during the tetanus. Our new findings on the cationic dependence of PTP are consistent with this hypothesis. 1) Several manuevers that decrease Na+-K+ pump activity, (decreasing [K+]o, replacing K+o with Rb+o or Li+o, or adding acetylstrophanthidin to Ringers), all increase the intratetanic rise and prolong the posttetanic decay of epp quantal content (m) and miniature epp frequency (fmepp). 2) Increasing [Ca2+]i or [Sr2+]o, but not [Mg2+]o, increases posttetanic fmepp in a graded fashion. 3) PTP of fmepp is still present after addition of Mn2+o, which blocks voltage dependent Ca2+ entry.
Origin and propagation of electrical slow waves in circular muscle of canine proximal colon
Authors: Smith T. K. , Reed J. B. , Sanders K. M.
Published: December 2017

Experiments to determine the site of slow-wave origin and the mechanism of propagation were performed on muscles of the canine proximal colon. Cells along the submucosal border of the circular layer had resting membrane potentials (RMP) averaging -78 mV, and slow waves, 40 mV in amplitude. The RMP of cells through the thickness of the circular layer decreased exponentially with distance from the submucosal border, such that RMPs of circular cells at the myenteric border were only -43 mV. Slow waves decreased in amplitude through the thickness such that slow waves could not be detected adjacent to the myenteric border. When a thin strip of muscle along the submucosal border was removed, slow waves were not recorded from the bulk of the circular layer and could not be evoked by acetylcholine. Slow waves were still present in the excised strip. Experiments to determine the rate of slow-wave propagation were also performed. Two cells were impaled, one at the submucosal surface, and another at some distance through the circular layer. Slow waves occurred nearly simultaneously at both sites. What latency was observed could be explained on the basis of electrotonic conduction. The results support the hypothesis that in the canine proximal colon slow waves are generated at the extreme submucosal surface of the circular layer. The bulk of the circular layer does not possess either pacemaker or regenerative mechanisms, and slow waves propagate passively toward the myenteric border. The cable properties of the circular muscle syncytium furnish a barrier to invasion of the longitudinal layer by the slow wave event.
Noradrenergic stimulation of mitochondriogenesis in brown adipocytes differentiating in culture
Authors: Nechad M. , Nedergaard J. , Cannon B.
Published: December 2017

The ability of adrenergic agents to promote the differentiation and especially the mitochondriogenesis of brown fat precursor cells, grown in culture, was investigated. These cells begin to differentiate during the days preceding confluence. We found here that, already during the early growth phase, the cultures (essentially precursor cells and preadipocytes at this stage) show increased cyclic AMP (cAMP) levels when acutely stimulated with norepinephrine (NE). The cultured cells were therefore chronically treated with NE up to the time of confluence, and their cytochrome-c oxidase activity was measured as an index of mitochondriogenesis. Chronic NE treatment resulted in an increased cytochrome-c oxidase activity of the cells at confluence. This effect was reproduced by selective activation of adenylate cyclase with cholera toxin, suggesting that the NE effect was exerted through an increase in cAMP. Ascorbate (added with NE as an antioxidant) had in itself a positive effect, both on final cell number and on cytochrome-c oxidase activity. It is concluded that NE, working through beta-adrenergic receptors, can stimulate mitochondriogenesis in brown fat cells through a direct effect on the cells, in accordance with the suggestion (based on in vivo experiments) that NE accelerates, rather than initiates, the differentiation process.
Molecular events in synaptogenesis: nerve-muscle adhesion and postsynaptic differentiation
Authors: Bloch R. J. , Pumplin D. W.
Published: December 2017

The clustering of acetylcholine receptors (AChR) in the postsynaptic membrane of newly innervated muscle fibers is one of the earliest events in the development of the vertebrate neuromuscular junction. Here, we describe two hypotheses that can account for AChR clustering in response to innervation. The "trophic factor" hypothesis proposes that the neuron releases a soluble factor that interacts with the muscle cell in a specific manner and that this interaction results in the local accumulation of AChR. The "contact and adhesion" hypothesis proposes that the binding of the nerve to the muscle cell surface is itself sufficient to induce AChR clustering, without the participation of soluble factors. We present a model for the molecular assembly of AChR clusters based on the contact and adhesion hypothesis. The model involves the sequential assembly of three distinct membrane domains. The first domain to form serves to attach microfilaments to the cytoplasmic surface of the muscle cell membrane at sites of muscle-nerve adhesion. The second domain to form is clathrin-coated membrane; it serves as a site of insertion of additional membrane elements, including AChR. Upon insertion of AChR into the cell surface, a membrane skeleton assembles by anchoring itself to the AChR. The skeleton, composed in part of actin and spectrin, binds and immobilizes significant numbers of AChR, thereby forming the third membrane domain of the AChR cluster. We make several predictions that should distinguish this model of AChR clustering from one that invokes soluble, trophic factors.
Alpha-actin and cytochrome c mRNAs in atrophied adult rat skeletal muscle
Authors: Babij P. , Booth F. W.
Published: December 2017

Specific complementary DNA (cDNA) hybridization probes were used to estimate the levels of alpha-actin and cytochrome c mRNAs and also 18S rRNA in three models of skeletal muscle atrophy. After 7 days of hindlimb suspension, or immobilization, or denervation, protein content decreased 26-32% in all muscles studied except suspended fast-twitch muscle, which lost only half as much protein. alpha-Actin mRNA content decreased 51-66% and cytochrome c mRNA content decreased 42-61% in slow- and fast-twitch muscles in all three models of atrophy. However, total RNA content did not show similar directional changes; RNA content decreased 27-44% in suspended and immobilized muscle but was unchanged in denervated fast-twitch muscle. The results were interpreted to suggest that loss of weight-bearing function of skeletal muscle is a major factor affecting the levels of alpha-actin and cytochrome c mRNAs during muscle atrophy.