George J. Siegel

Lithium transport across isolated frog skin epithelium.

SummaryTransepithelial Li+ influx was studied in the isolated epithelium from abdominal skin ofRana catesbeiana. With Na+-Ringers as inside medium and Li+-Ringers as outside medium, the Li+ influx across the epithelium was 15.6 μA/cm2. This influx was considerably reduced by removal of either Na+ or K+ from the inside bath or by the addition of ouabain or amiloride. Epithelial K+ or Na+ concentration was respectively lower in epithelia bathed in K+-free Ringers or Na+-free Ringers. In conditions of negligible Na+ transport, a 20mm Li+ gradient (out→in) produced across the short-circuited epithelium a Li+ influx of 11.8 μA/cm2 and a mean short-circuit current of 10.2 μA/cm2. The same Li+ gradient in the opposite direction produced a Li+ outflux of only 1.9 μA/cm2. With equal Li+ concentration (10.3 and 20.6mm) on both sides of the epithelium, plus Na+ in the inside solution only, a stable Li+-dependent short-circuit current was observed. Net Li+ movement (out→in) was also indirectly determined in the presence of an opposing Li+ gradient. Although Li+ does not substitute for Na+ as an activator of the (Na++K+)-ATPase from frog skin epithelium, Li+ influx appears to be related to Na+−K+ pump activity. It is proposed that the permeability of the “outer barrier” to Na+ and Li+ is regulated by the electrical gradient produced by electrogenic Na+−K+ pumps located in the membrane of the deeper epithelial cells.

Sodium-potassium-activated adenosine triphosphatase of brain microsomes: modification of sodium inhibition by diphenylhydantoins.

Effects of diphenylhydantoins on (Na(+) + K(+))-ATPase activity in rat and cat brain microsomes were studied. 5,5-diphenylhydantoin (DPH) in concentrations of 5-20 mug ml(-1) produces an apparent stimulation of the rat brain (Na(+) + K(+))-activated ATPase of 55-65% in media containing 50 mM Na(+), 0.15 mM K(+), 3 mM Mg(++), and 3 mM ATP. No effects are found on the Mg-ATPase. At constant K(+) levels of 0.05 mmole/liter and 0.15 mmole/liter, increasing the Na(+) concentration activates the enzyme similarly with and without DPH. However, Na(+) concentrations greater than 5 mmoles/liter and 10 mmoles/liter, respectively, which are inhibitory in these low K(+) media, produce less inhibition in the presence of DPH. In media containing 10 mM Na(+), the K(+) activation, on the other hand, is potentiated by DPH. In preparations from cat brain qualitatively similar results are obtained. No effect of DPH is seen on the inhibition produced by high K(+) in low Na(+) media. DPH produces an approximately constant apparent stimulation of 45% in the (Na(+) + K(+)) increments when these ions are varied simultaneously at a fixed ratio of 150 Na(+):1 K(+) with cat brain extracts. 5-(p-hydroxyphenyl)-5-phenylhydantoin (HPPH) has the same potency as DPH in reducing the Na(+) inhibition at high Na:K ratios. The hydantoins appear to act by decreasing the Na(+) inhibition that occurs at high Na:K ratios.

Nucleoside Triphosphate Phosphohydrolases

The pivotal role of ATP in energy metabolism is manifest in the frequency with which biological energy transducers are represented at the enzyme level as ATPases. Although some endergonic work processes may be coupled directly to electron transport or to electrochemical fluxes, energy transfer through ATP is apparently the most common mode.

Chapter 3 – Membrane Transport

Publisher Summary nPrimary active transporters are membrane proteins that enerxadgize many of the most basic neural functions. They transduce free energy from ATP hydrolysis into electrochemical energy that is stored in the transmembrane concentration gradixadents of Na+, K+, Ca2+ and protons. These energy stores are employed by membrane channel proteins for signaling, and by membrane secondary transporters that selectively concenxadtrate many other ions and molecules. Secondary active transxadporters depend on an ion gradient to transport their specific ligands uphill across membranes and subserve many neural functions such as packaging neurotransmitters into vesicles, terminating signals at synapses and transporting metabolites. The primary transporters discussed in this chapter belong to three distinct genomic superfamilies that differ markedly in structure and reaction mechanism. These are P-type, V0V1-type, and ABC-type. These superfamily members all catalyze reacxadtions with ATP that drive conformational cycles of the respective proteins to move substrates across membranes and “uphill” to higher concentrations.

Microsomal ouabain-binding factor in electroplax extracts.

Summary Lubrol WX extracts microsomal-bound ouabain-[3H] into the 50,000g supernatant fraction. The bound ouabain can be separated by chromatography with Sepharose and recovered in parallel fractions with recoverable adenosinetriphosphatase activity. Ouabain also binds to supernatant components in the Lubrol extracts and this bound ouabain has the same elution pattern as that bound to native microsomes prior to extraction with Lubrol. The proportion of bound ouabain that is recovered after chromatography is much greater than that of enzyme activity. Under these conditions, bound ouabain is a useful marker for enzyme extraction despite inactivation of the enzyme.