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L J Mullins

Publications and source records attributed to L J Mullins.

65 records · Page 4Linked to original sources
PubMed1967

Sodium extrusion by internally dialyzed squid axons.

A method has been developed which allows a length of electrically excitable squid axon to be internally dialyzed against a continuously flowing solution of defined composition. Tests showed that diffusional exchange of small molecules in the axoplasm surrounding the dialysis tube occurred with a half-time of 2-5 min, and that protein does not cross the wall of the dialysis tube. The composition of the dialysis medium was (mM): K isethionate 151, K aspartate 151, taurine 275, MgCI(2) 4-10, NaCl 80, KCN 2, EDTA 0.1, ATP 5-10, and phosphoarginine 0-10. The following measurements were made: resting Na influx 57 pmole/cm(2)sec (n = 8); resting potassium efflux 59 pmole/ cm(2)sec (n = 4); stimulated Na efflux 3.1 pmole/cm(2)imp (n = 9); stimulated K efflux 2.9 pmole/cm(2)imp (n = 3); resting Na efflux 48 pmole/cm(2)sec (n = 18); Q(10) Na efflux 2.2 (n = 5). Removal of ATP and phosphoarginine from the dialysis medium (n = 4) or external application of strophanthidin (n = 1) reversibly reduced Na efflux to 10-13 pmole/cm(2)sec. A general conclusion from the study is that dialyzed squid axons have relatively normal passive permeability properties and that a substantial fraction of the Na efflux is under metabolic control although the Na extrusion mechanism may not be working perfectly.

Adenosine Triphosphate↗
PubMed1967

Some factors influencing sodium extrusion by internally dialyzed squid axons.

Squid giant axons were internally dialyzed by a technique previously described. In an axon exposed to cyanide seawater for 1 hr and dialyzed with an ATP-free medium, the Na efflux had a mean value of 1.3 pmole/cm(2)sec when [Na](i) was 88 mM, in quantitative agreement with flux ratio calculations for a purely passive Na movement. When ATP at a concentration of 5-10 mM was supplied to the axoplasm by dialysis, Na efflux rose almost 30-fold, while if phosphoarginine, 10 mM, was supplied instead of ATP, the Na efflux rose only about 15-fold. The substitution of Li for Na in the seawater outside did not affect the Na efflux from an axon supplied with ATP, while a change to K-free Na seawater reduced the Na efflux to about one-half. When special means were used to free an axon of virtually all ADP, the response of the Na efflux to dialysis with phosphoarginine (PA) at 10 mM was very small (an increment of ca. 3 pmole/cm(2)sec) and it can be concluded that more than 96% of the Na efflux from an axon is fueled by ATP rather than PA. Measurements of [ATP] in the fluid flowing out of the dialysis tube when the [ATP] supplied was 5 mM made it possible to have a continuous measurement of ATP consumption by the axon. This averaged 43 pmole/cm(2)sec. The ATP content of axons was also measured and averaged 4.4 mM. Estimates were made of the activities of the following enzymes in axoplasm: ATPase, adenylate kinase, and arginine phosphokinase. Values are scaled to 13 degrees C.

Adenosine Triphosphatases↗
PubMed1967

Tracer and nontracer potassium fluxes in squid giant axons and the effects of changes in external potassium concentration and membrane potential.

The efflux of labeled and unlabeled potassium ions from the squid giant axon has been measured under a variety of experimental conditions. Axons soaked in sea water containing 42K ions lost radioactivity when placed in inactive sea water according to kinetics which indicate the presence of at least two cellular compartments. A rapidly equilibrating superficial compartment, probably the Schwann cell, was observed to elevate the specific activity of 42K lost from such axons to K-free sea water for a period of hours. The extra radioactive potassium loss from such axons during stimulation, however, was shown to have a specific activity identical within error to that measured in the axoplasm at the end of the experiment. The same was shown for the extra potassium loss occurring during passage of a steady depolarizing current. Axons placed in sea water with an elevated potassium ion concentration (50 mM) showed an increased potassium efflux that was in general agreement with the accompanying increase in membrane conductance. The efflux of potassium ions observed in 50 mM K sea water at different membrane potentials did not support the theory that the potassium fluxes obey the independence principle.

Action Potentials↗
PubMedDate not supplied

Changes in internal ionized Ca2+ and H+ in voltage clamped squid axons.

Squid giant axons were injected with aequorin and tetraethylammonium and were impaled with hydrogen ion sensitive, current and voltage electrodes. A newly designed horizontal microinjector was used to introduce the aequorin. It also served, simultaneously, as the current and voltage electrode for voltage clamping and as the reference for ion-sensitive microelectrode measurements. The axons were usually bathed in a solution containing 150 mM each of Na+, K+, and some inert cation, at either physiological or zero bath Ca2+ concentration [( Ca2+]o), and had ionic currents pharmacologically blocked. Voltage clamp pulses were repeatedly delivered to the extent necessary to induce a change in the aequorin light emission, a measure of axoplasmic ionized Ca2+ level, [( Ca2+]i). Alternatively, membrane potential was steadily held at values that represented deviations from the resting membrane potential observed at 150 mM [K+]o (i.e. approximately -15 mV). In the absence of [Ca2+]o a significant steady depolarization brought about by current flow increased [Ca2+]i (and acidified the axoplasm). Changes in internal hydrogen activity, [H+]i, induced by current flow from the internal Pt wire limited the extent to which valid measurements of [Ca2+]i could be made. However, there are effects on [Ca2+]i that can be ascribed to membrane potential. Thus, in the absence of [Ca2+]o, hyperpolarization can reduce [Ca2+]i, implying that a Ca2+ efflux mechanism is enhanced. It is also observed that [Ca2+]i is increased by depolarization. These results are consistent with the operation of an electrogenic mechanism that exchanges Na+ for Ca2+ in squid giant axon.

Aequorin↗
PubMedDate not supplied

Calcium entry in squid axons during voltage clamp pulses.

Squid giant axons were injected with aequorin and tetraethylammonium and were impaled with sodium ion sensitive, current and voltage electrodes. The axons were usually bathed in a solution of varying Ca2+ concentration ([Ca2+]o) containing 150mM each of Na+, K+ and an inert cation such as Li+, Tris or N-methylglucamine and had ionic currents pharmacologically blocked. Voltage clamp pulses were repeatedly delivered to the extent necessary to induce a change in the aequorin light emission, a measure of axoplasmic Ca2+ level, [Ca2+]i. The effect of membrane voltage on [Ca2+]i was found to depend on the concentration of internal Na+ ([Na+]i). Voltage clamp hyperpolarizing pulses were found to cause a reduction of [Ca2+]i. For depolarizing pulses a relationship between [Ca2+]i gain and [Na+]i indicates that Ca2+ entry is sigmoid with a half maximal response at 22 mM Na+. This Ca2+ entry is a steep function of [Na+]i suggesting that 4 Na+ ions are required to promote the influx of 1 Ca2+. There was little change in Ca2+ entry with depolarizing pulses when [Ca2+]o is varied from 1 to 10mM, while at 50mM [Ca2+]o calcium entry clearly increases suggesting an alternate pathway from that of Na+/Ca2+ exchange. This entry of Ca2+ at high [Ca2+]o, however, was not blocked by Cs+o. The results obtained lend further support to the notion that Na+/Ca2+ exchange in squid giant axon is sensitive to membrane voltage no matter whether this is applied as a constant change in membrane potential or as an intermittent one.

Aequorin↗