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Biomedical subjects

L J Mullins

Publications and source records attributed to L J Mullins.

At least 55 records · Page 3Linked to original sources
PubMed1977

Magnesium efflux in dialyzed squid axons.

The efflux of Mg++ from squid axons subject to internal solute control by dialysis is a function of ionized [Mg], [Na], [ATP], and [Na]o. The efflux of Mg++ from an axon with physiological concentrations of ATP, Na, and Mg inside into seawater is of the order of 2-4 pmol/cm2s but this efflux is strongly inhibited by increases in [Na]i, by decreases in [ATP]i, or by decreases in [Na]o. The efflux of Mg++ is largely independent of [Mg]i when ATP is at physiological levels, but in the absence of ATP reaches half the value of Mg efflux in be presence of ATP when [Mg]i is about 4 mM and [Na] 40 mM. Half-maximum responses to ATP occur at about 350 micronM ATP into seawater with Na either present or absent. The Mg efflux mechanism has many similarities to the Ca efflux system in squid axons especially with respect to the effects of ATP, Nao, and Na on the flux. The concentrations of free Mg and Ca in axoplasm differ, however, by a factor of 10(5) while the observed fluxes differ by a factor of 10(2).

Adenosine Triphosphate↗
PubMed1977

The control of ionized calcium in squid axons.

Measurements of the Ca content, [Ca](T), of freshly isolated squid axons show a value of 60 mumol/kg axoplasm. Axons in 3 mM Ca(Na) seawater show little change in Ca content over 4 h, while axons in 3 mM Ca(Na) seawater show little change in Ca content over 4 h, while axons in 10 mM Ca(Na) seawater show gains of 18 mumol/Ca/kgxh. In 10 Ca (Choline) seawater the gain is 2,400 mumol/kgxh. Using aequorin confined to a dialysis capillary in the center of an axon, one finds that [Ca](i) is in a steady state with 3 Ca (Na) seawater, and that both 10 Ca (Na) and 3 Ca (choline) seawater cause increases in [Ca](i). In 3 Ca (Na) seawater-3 Ca (choline) seawater mixtures, 180 mM [Na](0) (40 perecent Na) is as effective as 450 mM [Na](0) (100 percent Na) in maintaining a normal [Ca](1); lower [Na] causes an increase in [Ca](i). If axons are injected with the ATP-splitting enzyme apyrase, the resulting [Ca](1) is not loading with high [Ca](0) or low [Na](0) solutions. Depolarization of an axon with 100 mM K (Na) seawater leads to an increase in the steady-state level of [Ca](1) that is reversed upon returning the axon to normal seawater. Freshly isolated axons treated with either CN or FCCP to inhibit mitochondrial Ca buffering can still maintain a normal [Ca](i) in 1 Ca (Na) seawater.

Adenosine Triphosphate↗
PubMed1977

Intracellular calcium buffering capacity in isolated squid axons.

Changes in ionized calcium were studied in axons isolated from living squid by measuring absorbance of the Ca binding dye Arsenazo III using multiwavelength differential absorption spectroscopy. Absorption changes measured in situ were calibrated in vitro with media of ionic composition similar to axoplasm containing CaEGTA buffers. Calcium loads of 50-2,500 mumol/kg axoplasm were induced by microinjection, by stimulation in 112 mM Ca seawater, or by soaking in choline saline with 1-10 mM Ca. Over this range of calcium loading of intact axoplasm, the ionized calcium in the axoplasm rose about 0.6 nM/muM load. Similar loading in axons preteated with carbonyl cyanide 4- trifluoromethoxyphenylhydrazone (FCCP) to inhibit the mitochondrial proton gradient increased ionized calcium by 5-7 percent of the imposed load, i.e. 93-95 percent of the calcium load was buffered by a process insensitive to FCCP. This FCCP- insensitive buffer system was not saturated by the largest calcium loads imposed, indicating a capacity of at least several millimolar. Treatment of previously loaded axons with FCCP or apyrase plus cyanide produced rises in ionized calcium which could be correlated with the extent of the load. Analysis of results indicated that, whereas only 6 percent of the endogenous calcium in fresh axons is stored in the FCCP-sensitive (presumably mitochondrial) buffer system, about 30 percent of an imposed exogenous load in the range of 50-2,500 muM is taken up by this system.

Adenosine Triphosphate↗
PubMed1977

A mechanism for Na/Ca transport.

A model is developed which requires the binding of 4 Na+ to a carrier before a Ca binding site is induced on the opposite side of the membrane. Upon binding Ca, this carrier translocates Na and Ca. The existence of partially Na-loaded but nonmobile forms for the carrier (NaX, Na2X, Na3X) suffices to explain both the activating and the inhibitory effects of Na on the Ca transport reaction. Analytical expressions for Ca efflux and influx in terms of [Na]o, [Na]i, [Ca]o, [Ca]i, and Em are developed for the Na/Ca exchange system at equilibrium; these provide for a quantitative description of Ca fluxes. Under nonequilibrium conditions, appropriate modifications of the flux equations can be developed. These show a dependence of Ca efflux on [Ca]o and of Ca influx on [Ca]i. The large effect of internal ATP on Ca efflux and influx in squid axons, with no change in net Ca flux, can be understood on the single assumption that ATP changes the affinity of the carrier for Na at both faces of the membrane without providing an energy input to the transport reaction.

Adenosine Triphosphate↗
PubMed1976

Ionized calcium concentrations in squid axons.

Values for ionized [Ca] in squid axons were obtained by measuring the light emission from a 0.1-mul drop of aequorin confined to a plastic dialysis tube of 140-mum diameter located axially. Ionized Ca had a mean value of 20 x 10(-9) M as judged by the subsequent introduction of CaEGTA/EGTA buffer (ratio ca. 0.1) into the axoplasm, and light measurement on a second aequorin drop. Ionized Ca in axoplasma was also measured by introducing arsenazo dye into an axon by injection and measuring the Ca complex of such a dye by multichannel spectrophotometry. Values so obtained were ca. 50 x 10(-9) M as calibrated against CaEGTA/EGTA buffer mixtures. Wth a freshly isolated axon in 10 mM Ca seawater, the aequorin glow invariably increased with time; a seawater [Ca] of 2-3 mM allowed a steady state with respect to [Ca]. Replacement of Na+ in seawater with choline led to a large increase in light emission from aequorin. Li seawater partially reversed this change and the reintroduction of Na+ brought light levels back to their initial value. Stimulation at 60/s for 2-5 min produced an increase in aequorin glow about 0.1% of that represented by the known Ca influx, suggesting operationally the presence of substantial Ca buffering. Treatment of an axon with CN produced a very large increase in aequorin glow and in Ca arsenazo formation only if the external seawater contained Ca.

Aequorin↗
PubMed1976

Steady-state calcium fluxes: membrane versus mitochondrial control of ionized calcium in axoplasm.

Squid axons appear in a steady state with respect to ionized Ca when [Ca]o is ca. 3 mM and [Ca]i is ca. 30 nM. A membrane pump energized by the Na gradient across the membrane is capable of maintaining this ratio of ionized Ca if four Na enter per Ca extruded. Empirically, it is noted that if the difference between the electrochemical gradient for Na+ and that for Ca2+ is large and positive, Ca efflux is large; for a large but negative difference in the gradient, Ca influx is large. Net fluxes of Ca into the fiber are induced by Na-free solutions or stimulation. These are buffered in axoplasm principally by a high affinity Ca buffer. Mitochondria apparently do not contain large amounts of stored Ca as CN poisoning does not increase ionized Ca in axoplasm if the seawater is Ca-free.

Aequorin↗
PubMed1975

Sensitivity of calcium efflux from squid axons to changes in membrane potential.

Squid giant axons were internally dialyzed with a medium free of metabolic substrates but containing 45Ca buffered with EGTA to concentrations of free Ca++ in the range 0.01-230 muM. At (Ca)i of 1.0 muM OR GREATER, Ca efflux was in the range of 1-3 pmol/cm2 s, was dependent on (Na)o and (Ca)o, and was sensitive to membrane potential. At lower (Ca)i, the sensitivity of Ca efflux to membrane potential was greater. Hyperpolarization of the membrane increased, and depolarization decreased Ca efflux over the range of potentials studied (-20 to -100 mV). The maximum sensitivity of Ca efflux to membrane potential was of the order of an e-fold increase in Ca efflux for a 25-mV increase in Em; this sensitivity of Ca efflux to membrane potential was lost if (Na)o was removed and was greatly reduced when (Ca)i was increased to 230 muM.

Acetates↗
PubMed1975

Calcium and EDTA fluxes in dialyzed squid axons.

Ca efflux in dialyzed squid axons was measured with 45Ca as a function of internal ionized Ca in the range 0.005-10 muM. Internal Ca stores were depleted by treatment with CN and dialysis with media free of high energy compounds. The [Ca]iota was stabilized with millimolar concentrations of EDTA, EGTA, or DTPA. Nonspecific leak of chelated Ca was measured with [14C]-EDTA and found to be 0.02 pmol/cm2s/mM EDTA. Correction of the measured Ca efflux for this leak of chelated calcium was made when appropriate. Ca efflux was roughly linear with internal free Ca in the range 0.005-0.1 muM. Above 0.1 muM, efflux was less than proportional to concentration but did not saturate at the highest concentration studied. Ca efflux was reduced about 50% by replacement of external Na with Li at Caiota approximately 1 muM, but was insensitive to such replacement for Ca less than 0.1 muM. Ca efflux was insensitive to internal Mg in the range 0-4 mM, indicating that the Ca pump favors Ca over Mg by a factor of about 10(6). Ca efflux was reduced about 60% by increasing internal Na from 1 to 80 mM. This effect could represent weak interference of a Ca carrier by Na or a loss of driving force because of a reduction in ENa - Em occasioned by an increase in Naiota. A few measurements were made of Ca influx in intact and in dialyzed fibers. In both cases, Ca influx increased when external Na was replaced by Li.

Adenosine Triphosphate↗
PubMed1969

Potassium fluxes in dialyzed squid axons.

Measurements have been made of K influx in squid giant axons under internal solute control by dialysis. With [ATP](i) = 1 microM, [Na](i) = 0, K influx was 6 +/- 0.6 pmole/cm(2) sec; an increase to [ATP](i) = 4 mM gave an influx of 8 +/- 0.5 pmole/cm(2) sec, while [ATP](i) 4, [Na](i) 80 gave a K influx of 19 +/- 0.7 pmole/cm(2) sec (all measurements at approximately 16 degrees C). Strophanthidin (10 microM) in seawater quantitatively abolished the ATP-dependent increase in K influx. The concentration dependence of ATP-dependent K influx on [ATP](i), [Na](i), and [K](o) was measured; an [ATP](i) of 30 microM gave a K influx about half that at physiological concentrations (2-3 mM). About 7 mM [Na](i) yielded half the K influx found at 80 mM [Na](i). The ATP-dependent K influx responded linearly to [K](o) from 1-20 mM and was independent of whether Na, Li, or choline was the principal cation of seawater. Substances tested as possible energy sources for the K pump were acetyl phosphate, phosphoarginine, PEP, and d-ATP. None was effective except d-ATP and this substance gave 70% of the maximal flux only when phosphoarginine or PEP was also present.

Adenine Nucleotides↗
PubMed1968

Sodium fluxes in internally dialyzed squid axons.

The effects which alterations in the concentrations of internal sodium and high energy phosphate compounds had on the sodium influx and efflux of internally dialyzed squid axons were examined. Nine naturally occurring high energy phosphate compounds were ineffective in supporting significant sodium extrusion. These compounds were: AcP, PEP, G-3-P, ADP, AMP, GTP, CTP, PA, and UTP.(1) the compound d-ATP supported 25-50% of the normal sodium extrusion, while ATP supported 80-100%. The relation between internal ATP and sodium efflux was nonlinear, rising most steeply in the range 1 to 10 microM and more gradually in the range 10 to 10,000 microM. There was no evidence of saturation of efflux even at internal ATP concentrations of 10,000 microM. The relation between internal sodium and sodium efflux was linear in the range 2 to 240 mM. The presence of external strophanthidin (10 microM) changed the sodium efflux to about 8-12 pmoles/cm(2) sec regardless of the initial level of efflux; this changed level was not altered by subsequent dialysis with large concentrations of ATP. Sodium influx was reduced about 50 % by removal of either ATP or Na and about 70 % by removing both ATP and Na from inside the axon.

Adenine Nucleotides↗