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Biomedical subjects

D C Marcus

Publications and source records attributed to D C Marcus.

At least 55 records · Page 3Linked to original sources
PubMed1987

Sidedness of action of loop diuretics and ouabain on nonsensory cells of utricle: a micro-Ussing chamber for inner ear tissues.

It is known that nonsensory tissues of the utricle produce a lumen-positive transepithelial electrical potential difference (VT). This potential has been shown previously to be inhibited by ouabain and bumetanide applied to the bathing medium in vitro. In order to more fully characterize the origin of this potential we mounted the utricle as a flat sheet in a newly designed Ussing chamber and measured the VT and transepithelial resistance (RT) while perfusing the endolymphatic and perilymphatic surfaces independently with identical solutions. The aperture of the chamber was 1.5 X 10(-4) cm2. VT averaged 5.6 +/- 0.46 mV and RT was 24.0 +/- 2.43 omega X cm2 (n = 45). Ouabain and loop diuretics of the furosemide family were found to inhibit the VT only from the serosal side. The KI for ouabain was 7.63 X 10(-5) M. The loop diuretics tested inhibited the VT in the same order as in other tissues known to contain a Na/2 Cl/K cotransporter (KI: 2-benzylamino-4-cyclohexylsulfonyl-5-sulfamoylbenzolsulfonate++ + (BCSB), 1.72 X 10(-7) M; bumetanide, 1.10 X 10(-6) M; piretanide, 5.67 X 10(-6) M; furosemide, 4.14 X 10(-5) M). It is concluded that this tissue produces a lumen-positive VT (i) in the absence of a transepithelial chemical gradient; the generation of which is dependent upon the activity of (ii) Na,K-ATPase and (iii) a Na/2 Cl/K cotransporter; (iv) in the basolateral membranes of the nonsensory cells; (v) which is not depressed by luminal application of inhibitors of these transporters.

Animals↗
PubMed1987

Dependence of endocochlear potential on vascular pH.

The vasculature of the inner ear was perfused with simple salt solutions which were buffered with HCO3/CO2, PO4 or Hepes (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid). Replacement of HCO3/CO2 with either PO4 or Hepes at constant pH led to a rapid decline of about 15 mV in the endocochlear potential (EP) to a new steady-state level which could be maintained for over 30 min. This effect was reversible. Changes in [HCO3] over a wide range (nominally 0-100 mM) at constant CO2 tension produced only small (less than 3 mV) changes in the EP. However, the EP declined markedly when [HCO3] was maintained constant at 25 mM while CO2 tension was lowered. The response to increased CO2 was more complex. Additional experiments were performed in which intracellular pH was presumably altered by vascular perfusion of NH4 (alkalinization), or propionate (acidification). Perfusion of ammonium led to a strong decline of the EP (-38.2 +/- 2.5 S.D.) while propionate produced a small positive shift of about 3-4 mV. Acetazolamide (1 mM) decreased the EP by 7.6 +/- 2.7 mV and 14.8 +/- 4.6 mV in HCO3/CO2 and Hepes medium, respectively, after 10 min perfusion; this effect was poorly reversible. These results suggest that intracellular pH has a strong influence on the level of EP and further demonstrate that vascular [HCO3] and pH are not critical parameters for generation of the EP.

Acetazolamide↗
PubMed1987

Potassium secretion by nonsensory region of gerbil utricle in vitro.

The isolated nonsensory region of the gerbil utricle in vitro produced a lumen-positive transepithelial electrical potential difference (VT) of +5.7 mV and a luminal fluid containing 106 mM K when bathed in mammalian Ringer solution (5 mM K and 150 mM Na). The lumen of this region was perfused in vitro with K-free solution and the luminal [K], VT, and transepithelial resistance (RT) were measured before and following perfusion under control conditions and after addition of bumetanide (0.1 mM) or ouabain (1 mM) to the bath. The perfusate contained a reduced [Ca], since the average value of utricular endolymph in vivo (0.28 +/- 0.03 mM) measured with Ca-selective microelectrodes was 38% of that in perilymph. Under control conditions, the luminal [K] initially increased at a rate of 2.13 mumol X cm-2 X h-1 after perfusion; net secretion continued until the luminal [K] returned to its preperfusion level. This flux rate corresponds to 57 microA/cm2. The "equivalent short-circuit current" (Equiv. Isc; VT/RT) was found to average 61 microA/cm2. Both K secretion and VT were fully inhibited by bumetanide and by ouabain. Luminal application of Ba (5 mM) in K-free solution had no effect on the initial rate of K secretion, but did prevent full recovery of luminal [K] to the control level. These results are the first estimates of K secretion by the nonsensory cells of the utricle and are the first to directly demonstrate inhibition of K secretion in the inner ear by bumetanide and in the nonsensory tissue of the utricle by ouabain.

Animals↗
PubMed1987

Effect of iodoacetic acid upon cochlear potentials.

Lotz et al. reported that perilymphatic application of 5 X 10(-3) M iodoacetic acid (IAA) in the guinea pig does not influence the first-order cochlear microphonics (CM1) under aerobic conditions. However, in ischemia the rate of decline of the second-order microphonics (CM II, also called postmortem CM) was significantly increased by IAA. The authors concluded that glycolysis plays no role in maintaining the CMI, but that it is responsible for supporting the CMII. In carefully controlled experiments we found that in the respiring guinea pig, perilymphatic application of 5 X 10(-3) M IAA produced a rapid and pronounced effect upon both the endolymphatic potential and the CM. In particular, the CM dropped to less than 0.5% of its initial level within 40 min, due to IAA, whereas it took 120 min to drop to the same level in total ischemia (without IAA). We therefore reject the above-mentioned proposition that IAA is ineffective upon cochlear potentials under aerobic conditions; moreover, we find that even under aerobic conditions, the CM drops well below the usual CM II level substantially faster than under anaerobic conditions (without IAA). Other important findings, including an anoxia-sensitive negative component of the endolymphatic potential due to severe intoxication with IAA, and the effects of pretreatment of the organ of Corti with low concentrations of IAA upon the CM II are discussed.

Aerobiosis↗
PubMed1986

Membrane transport parameters in frog corneal epithelium measured using impedance analysis techniques.

Active Cl- transport in bullfrog corneal epithelium was studied using transepithelial impedance analysis methods, and direct-current (DC) measurements of membrane voltages and resistance ratios. The technique allows the estimation of the apical and basolateral membrane conductances, and the paracellular conductance, and does not rely on the use of membrane conductance-altering agents to obtain these measurements as was requisite in earlier DC equivalent-circuit analysis studies. In addition, the analysis results in estimates of the apical and basolateral membrane capacitances, and allows resolution of the paracellular conductance into properties of the tight junctions and lateral spaces. Membrane capacitances (proportional to areas) were used to estimate the specific conductances of the apical and basolateral membranes, as well as to evaluate coupling between the cell layers. We confirm results obtained from earlier studies: apical membrane conductance is proportional to the rate of active Cl- transport and is highly Cl- selective; intracellular Cl- activity is above electrochemical equilibrium, thereby providing a net driving force for apical membrane Cl- exit; the paracellular conductance is comparable to the transcellular conductance. We also found that: the paracellular conductance is composed of the series combination of the junctional conductance and a nonnegligible lateral space resistance; a small K+ conductance reported in the apical membrane may result from Cl- channels possessing a finite permeability to K+; the basolateral membrane areas is 36 times greater than the apical membrane area which is consistent with the notion of electrical coupling between the five to six cell layers of the epithelium; the specific conductance of the basolateral membrane is many times lower than that of the apical membrane; the net transport of Cl- is modulated primarily by changes in the conductance of the apical membrane and not by changes in the net electrochemical gradient resulting from opposite changes in the electrical and chemical gradients; the conductance of the basolateral membrane does not change with transport which implies that the net driving force for K+ exit increases with transport, possibly due to an increase in the intracellular K+ activity.

Animals↗
PubMed1986

Direct measurement of longitudinal endolymph flow rate in the guinea pig cochlea.

The rate of longitudinal endolymph flow in the guinea pig cochlea has been measured with a novel tracer technique. The tracer we utilized was the tetramethylammonium (TMA) ion, the movement of which was monitored by ion-sensitive microelectrodes. Extremely small amounts of tracer were required as the electrodes could readily detect TMA concentrations in endolymph as low as 10 microM. TMA was introduced into scala media in the form of a small bolus, varying from 2-20 nl in volume. To examine whether longitudinal flow affects the dispersion of TMA in endolymph, we compared the characteristics of TMA spread to turn I following injection into turn II, with those of TMA spread to turn II following injection into turn I. The comparison of these data with an analytical model combining the processes of diffusion and volume flow demonstrates that the spread of tracer is dominated by passive diffusion processes with very little contribution from longitudinal endolymph flow. The rate of longitudinal endolymph flow between turn I and turn II was estimated to be less than 0.01 mm/min directed towards the basal turn. This value is considerably lower than recently published estimates using other techniques.

Cochlea↗
PubMed1986

Transepithelial electrical potential of nonsensory region of gerbil utricle in vitro.

Transepithelial electrical potential difference (VT) was measured across the vestibular labyrinth of the inner ear in vitro by puncturing the epithelial wall of the utricle with a glass microelectrode. A region of nonsensory cells of the utricle was isolated from the sensory regions by introducing columns of liquid Sylgard 184. Under control conditions, the VT of this region was +7.5 +/- 0.3 mV (means +/- SE), lumen positive. This potential difference was rapidly reduced by either 1 mM ouabain, 10-100 microM bumetanide, 0.5-5.0 mM Ba (in the bathing solution), or cooling, but not by the disulfonic stilbene, 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid. Changes in VT due to reductions of Cl or Na or to increases of K in the bathing solution in exchange for presumably impermeant ions were observed in this region and were compared with those in a preparation in which the insulating seals were absent. The K-induced voltage change was significantly higher in the unblocked preparation, a finding consistent with a high K permeability of the sensory cells. The voltage change due to reduction of Cl was not inhibited by Cl channel blockers (9-anthracenecarboxylate and diphenylamine-2-carboxylate) in the bathing solution. These results represent the first direct demonstration that the nonsensory cells of the utricle produce a lumen-positive active-transport potential and characterize some of the properties of the cell membranes in terms of their pharmacological sensitivities and net voltage responses to changes in the bathing medium ions Na, K, and Cl.

Animals↗
PubMed1985

Effects of barium and ion substitutions in artificial blood on endocochlear potential.

Previously, a qualitative assessment was made (Marcus, D.C. (1984): Am. J. Physiol. 247, C240-C246) of the ion-selective properties of the cells bounding the cochlear duct by observing the effects of ion substitutions in the perilymph on the transepithelial potential difference (endocochlear potential; EP). Contributions by the marginal cells of the stria vascularis to the observed changes in the EP may have been masked, however, due to their 'isolation' from the perilymph by a continuous layer of basal cells. Since the ionic milieu of the basolateral membranes of the marginal cells is controlled more directly by the blood supply than by the perilymph, we report here on the effects of ion substitutions via vascular perfusion. Elevated K (substituted for Na or N-methyl-D-glucamine; NMDG) or Ba caused marked depression of the EP. Decreased Na or Cl (replaced by NMDG and gluconate, respectively) also depressed the EP. These changes in the EP were distinctly different from those observed previously by perilymphatic perfusion, and were interpreted in terms of a modified model of the strial marginal cells.

Barium↗
PubMed1984

Prolonged maintenance of endocochlear potential by vascular perfusion with media devoid of oxygen carriers.

A method is described for maintaining the cochlear potentials of the guinea pig via arterial perfusion of the surviving inner ear with an artificial medium devoid of oxygen carriers or oncotic agents. The endocochlear potential (EP) can be maintained at a normal level for periods in excess of 5 h; the responses of the EP to anoxia and to furosemide closely approximate those seen in the intact animal. This preparation may represent a simplified method for carrying out selected arterial perfusion experiments in the surviving inner ear.

Animals↗
PubMed1984

Support of cochlear metabolic and ion transport processes solely by perilymphatic perfusion.

Morphologic considerations would seem to suggest that the cochlear duct could not be maintained in a fully functional state in the absence of a blood supply. We found, however, that perilymphatic perfusion could be used as a substitute for the normal vascular circulation. The criteria used to determine cochlear function included (1) normal endocochlear potential, (2) normal net secretory flux of rubidium (as a tracer for K), and (3) normal levels of ATP in both the organ of Corti and the stria vascularis. All criteria were satisfied by our perfusion regimen.

Adenosine Triphosphate↗
PubMed1984

Characterization of potassium permeability of cochlear duct by perilymphatic perfusion of barium.

The relative transepithelial "permeabilities" of the cochlear duct to K, Na, and Cl were investigated so as to identify the K-selective tissues and to determine the cellular origin of this selectivity. Single-ion substitutions were made for K, Na, and Cl with the impermeant species N-methyl-D-glucamine (NMDG) for K and Na and gluconate or sulfate for Cl in perilymph. Transepithelial potential changes were relatively slow and small for Na and Cl substitutions. However, either K for Na or K for NMDG substitutions demonstrated a pronounced K selectivity (rapid changes of electrical potential) of only the sensory-cell tissue (organ of Corti). The response to the K for Na substitution was most clearly seen after electrogenic K transport was inhibited by ischemia while the sensory cells were metabolically sustained via perilymphatic perfusion. Under this condition, perfusion of a medium containing 154 mM K gluconate reduced the negative potential (typically -25 to -40 mV) to within a few millivolts of zero. In a control medium, perilymphatic barium (0.5-5 mM) produced qualitatively similar effects, suggesting that this K selectivity is localized primarily at the basolateral membrane of the sensory cells rather than at the junctional complexes.

Animals↗
PubMed1983

Response of cochlear potentials to presumed alterations of ionic conductance: endolymphatic perfusion of barium, valinomycin and nystatin.

Two models ('single-pump' and 'two-pump') of transepithelial potassium movement by the marginal cells of the stria vascularis have been proposed in the literature. Their validity was considered by exposing the endolymphatic (luminal) surface to agents (barium, valinomycin and nystatin) which are known to alter specific cellular membrane conductances in other tissues. This was accomplished by the use either of injections or of a relatively satisfactory technique for perfusion of scala media, which is described. Injection of barium caused the endocochlear potential (EP) to increase in normal animals and had no effect on the EP of deaf, Waltzing guinea pigs. Perfusion of the ionophores caused a decline in the EP in both normal and Waltzing guinea pigs. Only the 'two-pump' model (Na/K-ATPase-mediated cation pump on the basolateral membrane and rheogenic K transporter at the luminal membrane) is consistent with the results. The cellular heterogeneity of the cochlear duct, however, introduces a measure of uncertainty into this interpretation.

Animals↗
PubMed1982

Comparison of the non-adrenergic action of phentolamine with that of vanadate on cochlear function.

Two drugs, which upon superficial examination appeared to be acting on common processes, have been found upon closer investigation to act by quite different means. Both act primarily at the organ of Corti, causing a pronounced increase of the endocochlear potential and a depression of the cochlear microphonic (CM). These effects are accompanied by the elimination of a negative component of the EP; however, it was found that these three effects are produced by phentolamine in scala media (or, more slowly, in scala tympani) but by vanadate only in scala tympani. This difference in locus of action isd manifested further by different changes of the summating potential (SP): phentolamine has little effect on the magnitude of SP-, while vanadate leads to an elevated SP-. In spite of this difference in the 'zeroth order harmonic', the second harmonic of the CM is depressed by both agents. It is argued that phentolamine may act either by blocking the acoustically-modulated ion channels in the luminal membranes of the hair cells or by inducing a large, non-selective, paracellular conductance in the organ of Corti. The present results, in conjunction with our previous results (Marcus, D.C., DeMott, J.E., Kobayashi, T., Ge, X.-X. and Thalmann, R. (1981): Hearing Res. 5, 231-243), are further interpreted as suggesting that vanadate may initially act by depolarizing the hair cells.

Adrenergic alpha-Antagonists↗
PubMed1981

Changes in cation contents of stria vascularis with ouabain and potassium-free perfusion.

Perfusion of the perilymphatic space of guinea pig cochleae with K-free medium leads to a gradual decline of the endocochlear potential (EP) over 30-50 min to a negative value (mean: -12 mV). The input resistance of scala media does not decrease during this time. The ATP and K content of the stria vascularis are reduced by similar amounts (26 and 34%, respectively) during this period. Perfusion of 1 mM ouabain produces a different pattern of response: strial ATP remains normal while strial K content is strongly reduced (by 77%). Strial Na rises in a complementary way to the K loss. These results demonstrate that a reduction of the K concentration of the perilymph leads to an inhibition of the generator of the positive component of the EP rather than to a general increase of cochlear duct membrane conductance. In addition, they suggest, in concert with other considerations (such as the slower rate of decline of the EP during K-free vascular perfusion (Wada, J., Kambayashi, J., Marcus, D.C. and Thalmann, R (1979): Arch. Otorhinolaryngol. 225, 79-81)), that the mode of action may be different from that of ouabain. In spite of the lack of teleological support, we offer the hypothesis that the strial generator of the EP may primarily utilize K from perilymph and that vascular K may not have access to the generator.

Adenosine Triphosphate↗
PubMed1981

Specificity of action of vanadate to the organ of corti.

Although vanadate strongly inhibits Na/K-ATPase activity of the stria vascularis in vitro, it initially causes no depression of the ouabain-sensitive endocochlear potential (EP) when perfused perilymphatically or via the vasculature. However, when the perilymph of scala tympani is replaced with artificial media containing 0.1 to 1 mM vanadate, there is a large (about 17 mV) increase in the EP of the second cochlear turn. Further experiments showed that the cochlear microphonics declined during the time in which the EP increased, and that the response of these two potentials to vanadate is greater in the second turn than in the first. Injection of 50 n1 of 1 mM vanadate (in artificial endolymph) into the endolymphatic space of the second turn caused no increase in the EP. These results support the notion that the early effects of vanadate are on the contra-luminal membranes of cells of the organ of Corti rather than on the stria vascularis. By superimposing anoxia or furosemide (i.v.) upon vanadate intoxication, we determined that the initial increase of the compound EP due to vanadate alone was due to a reduction in magnitude of the negative component of the EP. It is argued that of the three prevalent theories concerning the generation of the negative EP, the data tend to support the hypothesis that the intracellular potential of the hair cells gives rise to the negative EP.

Adenosine Triphosphatases↗
PubMed1979

Vascular perfusion of the cochlea: effect of potassium-free and rubidium-substituted media.

By means of vascular perfusion of the cochlea with K+-free synthetic blood, it is possible to maintain the endocochlear potential (EP) at normal or supernormal levels for periods of 20-42 min. The subsequent decline of the EP is comparatively slow. This is in contrast to the immediate onset and rapid decline of the EP during perilymphatic perfusion with K+-free media. Possible implications of these data concerning the mode of generation of the EP are briefly discussed. It is further shown, that synthetic blood, in which K+ is replaced by Rb+, maintains the EP at normal or near normal levels for 2 h.

Cochlea↗