Weak acid accumulation in the serosal extracellular compartment of the frog gastric mucosa

The dimethyloxazolidine dione distribution in the extracellular compartments of the frog gastric mucosa was analyzed by washout kinetics. The volumes of the two extracellular compartments, serosal and mucosal, were estimated by inulin washout as $\text{0.435 ± 0.019}$ and $\text{0.176 ± 0.018 μl/μl}$ tissue water, respectively. In the serosal extracellular space, significant dimethyloxazolidine dione accumulations of $\text{2.63 ± 0.25}$, $\text{2.28 ± 0.16}$, and $\text{1.86 ± 0.08}$ times that of the bathing media were found for bathing solutions with pH values of 6.9, 7.4, and 7.9 respectively. A high pH of the serosal extracellular fluid by itself could not account for the high values of dimethyloxazolidine dione accumulation. A difference in the total dimethyloxazolidine dione accumulation requires: (a) the existence of differences in the pH values and also the existence of a difference in the diffusion coefficient of the two forms of dimethyloxazolidine dione; or (b), a binding of one of the two forms, i.e., binding of dimethyloxazolidine dione form by fixed charges.     

Stereochemical course of the biosynthesis of 1-aminocyclopropane-1-carboxylic acid. I. Role of the asymmetric sulfonium pole and the α-amino acid center

The substrate stereospecificity of 1-aminocyclopropane-1-carboxylic acid synthase, a pyridoxal phosphate-containing enzyme, from the pericarp tissue of $\text{Lycopersicon esculentum}$ (tomatoes) was studied using the various stereoisomers of $\text{S}$-adenosylmethionine (AdoMet) at both the sulfonium pole and the amino acid center. The data indicate that only the naturally occurring isomer (?)Ado-L-Met acts as substrate (Km = 20±5 μM). Both (±)Ado-D-Met and (+)Ado-L-Met were inactive as substrates. The (+)Ado-L-Met (Ki = 15±5 μM) was found to be a potent inhibitor of ACC synthase whereas (±)Ado-D-Met (Ki = 70±20 μM) was less active as an inhibitor. This active isomer has the ($\text{S}$) configuration at both the sulfur and the α-carbon of the amino acid portion of AdoMet.     

Cellular pH and water permeability control in frog urinary bladder a possible action on the water pathway

Mucosal acidification (from pH 8.1 to 6.0) reversibly inhibited the hydroosmotic responses to oxytocin, cyclic AMP and 8-bromo-cyclic AMP in frog urinary bladder. These inhibitory effects were only observed in the presence of a permeant buffer in the apical medium and could also be elicited by CO₂ bubbling, even when the mucosal pH was clamped at 8.1. Acid pH reduced the oxytocin-induced net water flux faster than norepinephrine or oxytocin removal and the difference was especially important at low temperature. The time course of recovery from acid pH inhibition was, at 20°C, similar to that of the hormonal action, but when the medium temperature was reduced to 6–7°C, the recovery from acid pH inhibition paradoxically became faster while the oxytocin action was markedly slowed down ($\text{t}{}_{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$ of changes in net water fluxes (expressed in min): oxytocin addition at 20°C, $\text{6.2 ± 0.9}$; at 6°C, $\text{24 ± 3}$; oxytocin removal at 20°C, $\text{4.7 ± 0.8}$; at 6°C, $\text{22 ± 3}$; pH inhibition at 20°C, $\text{2.6 ± 0.2}$; at 6°C $\text{2.5 ± 0.2}$; recovery from pH 6 at 20°C, $\text{6.5 ± 0.9}$; at 6°C, $\text{2.7 ± 0.3}$). These results can be explained by accepting two main loci sensitive to medium acidification: (1) the cyclase system and (2) an intracellular, temperature-independent, post-cyclic AMP site. The fact that the intramembranous particle aggregates associated with the oxytocin-induced water permeability increase did not disappear after the flow inhibition by acid pH at low temperature suggests that the second effect could be located at the water channel itself.     

A new assay system of phospholipid exchange activities using concanavalin a in the separation of donor and acceptor liposomes

A new assay system of phospholipid exchange activities is described. The exchange activities were quantitated by measuring the stimulation of phospholipid transfer between two separate populations of liposomes, which contained, as the major constituents, phosphatidylcholine, phosphatidylethanolamine, phosphatidylinositol, sphingomyelin, and cholesterol in molar ratios of 6: 2: 1: 1: 5. One population of the liposomes was made reactive to concanavalin A by the incorporation of 1.8 mol% $\text{α-}\text{d}\text{-mannosyl}\text{-(1 → 3)-α-}\text{d}\text{- mannosyl}\text{-sn-}\text{1,2-diglyceride}$ from Micrococcus lysodeikticus. The concanavalin A-reactive liposomes, a phospholipid donor, were doubly labelled with [6-³H]galactosylglucosyl ceramide and that class of ³²P-labelled phospholipids whose exchange was being measured. The ³H-labelled glycolipid served as a non-exchangeable reference marker. The other population of the liposomes, a phospholipid acceptor, was concanavalin A nonreactive. These two populations of liposomes were incubated with the cytosol protein of rat liver in a total volume of 0.2 ml.After the incubation, two different procedures were used to separate the two liposomal populations. In one procedure concanavalin A was added to agglutinate the reactive liposomes; the flocculated lectin-liposome complex was separated from the non-reactive liposomes by brief centrifugation. In the other procedure the reactive liposomes were trapped by binding to concanavalin A covalently coupled to Sepharose 2B; the complex was separated from the nonreactive liposomes by filtration through a filter paper under suction. In both assay procedures the amount of phospholipid transferred from the donor to the acceptor liposomes was calculated from the decrease of ³²P/³H ratio of the concanavalin A-reactive liposomes during the incubation. By the assay system it is possible to determine phosphatidylcholine and phosphatidylinositol exchange activities in 100 μg of rat liver cytosol protein.     

Amphotericin B-induced sodium transport and water flow across rabbit corneal epithelium

We have determined fluid translocation across the cellular layers lining the cornea by measuring changes in corneal transparency. The loss of 1.3 μ1/cm² fluid from the stroma causes an increase of +1% in transparency. Amphotericin B ($\text{2 · 10}{}^{-6}\text{M}$) when added to the tear side (=mucosal side) of the epithelium causes a rapid increase in potential difference of $\text{12.3 ± 0.7}\text{mV}\text{(}\text{mean}\text{±}\text{S.E.}\text{, n=6}$) followed by a slower increase of $\text{18.6 ± 1.5}\text{mV}$. The electrical resistance is reduced from $\text{3.2 ± 0.3}\text{k}\text{Ω ·}\text{cm}{}^2\text{to}\text{0.6 ± 0.1}\text{k}\text{Ω ·}\text{cm}{}^2$. The resulting increase in calculated short circuit current is accompanied by a decrease in transparency at a rate of $\text{3.6 ± 1.0\%}\text{per h}$, corresponding to an uptake of fluid by the cornea of $\text{4.7 μ}\text{l}\text{·}\text{cm}{}^{-2}\text{·}\text{h}{}^{-1}$. Replacement of the fluid bathing the endothelial side of the cornea, in order to prevent water movement from the aqueous compartment into the stroma, did not significantly alter this uptake of fluid. Thus the epithelial fluid transport which is reported to be normally slightly secretory, becomes absorptive in the presence of amphotericin B. Serosal hypertonicity (20 mM mannitol) increases the water influx into the cornea induced by amphotericin B. These results indicate that amphotericin B induces sodium-selective channels in the epithelium leading to an accumulation of NaCl and water in the stromal layer of the cornea. Ouabain reduces the potential and calculated short circuit current in epithelia pretreated with amphotericin B. Following addition of ouabain, the NaCl and water accumulated in the stroma leak away resulting in a transient increase in transparency. Finally, a model is proposed that includes a stromal compartment involved in fluid transport and that agrees with the results presented here.     

An ultrastructural examination of everted rat jejunum

Male, albino, Sprague-Dawley rats were sacrificed by cervical separation. Segments of jejunum were excised, everted and examined with the electron microscope. Examination of tissue fixed immediately after eversion revealed the following changes as compared to non-everted segments fixed $\text{in}$$\text{situ}$ and $\text{in}$$\text{vitro}$: 1) an increase in the length of microvilli from (mean ± S. E.) 0.991 ± 0.011μ for normal tissue to 1.389 ± 0.023μ for everted tissue, 2) an increase in width of microvilli from (mean ± S. E.) 0.089 ± 0.001μ for normal tissue to 0.097 ± 0.001μ for everted tissue, 3) an increase in length and number of lateral membrane interdigitations, and 4) the appearance of intercellular “lakes” in the lateral spaces. The above changes are in those structures hypothesized to be involved with salt and water transport across epithelia and may reflect altered transport rates $\text{in}$$\text{vitro}$ as compared to $\text{in}$$\text{vivo}$.     

The role of microfilaments and of microtubules in taurocholate uptake by isolated rat liver cells

The role of microfilaments and microtubules on bile salt transport was studied by investigating the influence of a microfilament and a microtubule inhibitor, cytochalasin B and colchicine, respectively, on taurocholate uptake by isolated hepatocytes in vitro. Hepatocytes were prepared by the enzyme perfusion method and [¹⁴C]taurocholate uptake velocity was determined by a filtration assay. Taurocholate uptake obeyed Michaelis-Menten kinetics, maximal uptake velocity and apparent half-saturation constants averaging $\text{0.87 ±}\text{SD}\text{0.05}\text{nmol}\text{· s}{}^{\mathrm{?1}}\text{· 10}{}^{\mathrm{?6}}\text{cells}$ and $\text{10.9 ± 1.8 μ}\text{M}$, respectively. Cytochalasin B ($\text{4.2–420 μ}\text{M}$) inhibited taurocholate uptake in a competitive fashion; $\text{K}{}_{\mathrm{<mtext>i</mtext>}}$ being $\text{33 ± 7 μ}\text{M}$. At concentrations above 100 μM the compound decreased ³⁶Cl membrane potential and intracellular K⁺ concentration. Other parameters of cell viability were not affected by cytochalasin B. Colchicine (0.1–1.0 mM), by contrast, inhibited taurocholate uptake non-competitively, $\text{K}{}_{\mathrm{<mtext>i</mtext>}}$ being $\text{0.47 ± 0.07}\text{mM}$. The inhibition brought about by colchicine was considerably smaller than that induced by cytochalasin B. None of the parameters of cell viability tested was affected by colchicine. These results suggest that microfilaments may be involved in the carrier-mediated hepatocellular transport of bile salts. This could, at least in part, account for cytochalasin B-induced cholestasis. The contribution of the microtubular system, if any, is less important quantitatively. The mechanisms whereby these two components of the cytoskeleton partake in bile salt transport remain to be elucidated.     

Immunologic response to protein immobilized on the surface of liposomes via covalent azo-bonding

A new method for immobilizing protein on the surface of liposomes is described. Inclusion of $\text{N-(p-}\text{aminophenyl)}$stearylamide in the lipid composition of vesicles resulted In liposomes that could be ‘activated’ by diazotization with NaNO₂/HCl, and subsequently coupled with protein. Using this method $\text{39.7 ? 7.5 μ}\text{g}$ egg albumin / μmol phospholipid has been coupled to multilamellar vesicles composed of phosphatidylcholine, cholesterol, and $\text{N-(p-}\text{aminophenyl}\text{)}$stearylamide in a molar ratio of 15:7.5:1.1. Furthermore, when the immunologic response of mice to egg albumin that was encapsulated in, nonspecifically adsorbed, or covalently linked to liposomes was investigated, only the covalent protein-liposome conjugates elicited pronounced and sustained elevations in antibody titers. These results suggest that the immunoadjuvant effects of liposomes can be maximized by covalently linking protein antigens to their surface.     

Asymmetric or symmetric? Cytosolic modulation of human erythrocyte hexose transfer

(1) The Michaelis-Menten parameters for hexose transfer in erythroctes, erythrocyte ghosts and inside-out vesicles at 20°C were determined using the light scattering method of Sen and Widdas ((1962) J. Physiol. 160, 392–403). (2) The external $\text{K}{}_{\mathrm{<mtext>m</mtext>}}$ for infinite-cis exit of d-glucose in cells and ghosts is $\text{3.6 ± 0.5}\text{mM}$. (3) Dilution of cellular solute (up to × 90 dilution) by lysing and resealing cells in varying volumes of lysate is without effect on the $\text{V}{}_{\mathrm{<mtext>m</mtext>}}$ for net d-glucose exit. The $\text{K}{}_{\mathrm{<mtext>m</mtext>}}$ for net exit, however, falls from $\text{32.4 ± 3.7}\text{mM}$ in intact cells to $\text{12.9 ± 2.3}\text{mM}$ in ghosts. This effect is reversible. (4) Infinite-cis net d-glucose uptake measurements in cells and ghosts reveal the presence of a low $\text{K}{}_{\mathrm{<mtext>m</mtext>}}$, high affinity internal site of $\text{5.9 ± 0.8}\text{mM}$. The $\text{V}{}_{\mathrm{<mtext>m</mtext>}}$ for net glucose entry increases from $\text{23.2 ± 3.7}\text{mmol/l per min}$ in intact cells to $\text{55.4 ± 6.3}\text{mmol/l per min}$ in ghosts. (5) The external $\text{K}{}_{\mathrm{<mtext>m</mtext>}}$ for infinite-cisd-glucose exit in inside-out vesicles is $\text{6.8 ± 2.7}\text{mM}$. The kinetics of zero-transd-glucose exit from inside-out vesicles are changed markedly when cellular solute (obtained by lysis of intact cells) is applied to either surface of inside-out vesicles. When solute is present externally, the $\text{K}{}_{\mathrm{<mtext>m</mtext>}}$ and $\text{V}{}_{\mathrm{<mtext>max</mtext>}}$ for zero-trans exit are decreased by up to 10-fold. When solute is present at the interior of inside-out vesicles, $\text{V}{}_{\mathrm{<mtext>max</mtext>}}$ for zero-trans exit is reduced; $\text{K}{}_{\mathrm{<mtext>m</mtext>}}$ for exit is unaffected. In the nominal absence of cell solute, transfer is symmetric in inside-out vesicles. The orientation of transporter in the bilayer is unaffected by the vesiculation procedure. (6) External application of cellular solute to ghosts reduces $\text{V}{}_{\mathrm{<mtext>max</mtext>}}$ for d-glucose exit but is without effect on the external $\text{K}{}_{\mathrm{<mtext>m</mtext>}}$ for infinite-cis exit. (7) The inhibitory potency of cell lysate on hexose transfer is lost following dialysis indicating that the factors responsible for transfer modulation are low molecular weight species. (8) We consider the hexose transfer in human erythrocytes is intrinsically symmetric and that asymmetry of transfer is conferred by interaction of the system with low molecular weight cytosolic factors.     

The content of water and potassium in fat cells

The distribution spaces at equilibrium for ³H₂O, [¹⁴C]urea and $\text{3-O-[}{}^{14}\text{C]-}\text{methylglucose}$ were measured in white fat cells using centrifugation through silicone oil at 2500 × g; no significant differences were observed. l-[¹⁴C] Glucose added immediately before the centrifugation was used as a marker for the extracellular water space. The calculated intracellular water content of the cells after the centrifugation through oil (e.g. ³H₂O space minus l-[¹⁴C] glucose space) is an unbiased measure of the water content of the cells in suspension as judged by the following criteria: (1) The intracellular distribution space for $\text{3-O-[}{}^{14}\text{C}\text{]}$methylglucose at equilibrium (methylglucose space minus l-glucose space) was not different from that calculated from a methylglucose wash-out curve. (2) The intracellular content of l-[¹⁴C]glucose (half time of efflux about 60 min) in cells preloaded during incubation of the tissue with collagenase was not different in cells recovered by (a) centrifugation through oil at 2500 × g, (b) centrifugation through oil at 600 × g, (c) centrifugation at 600 × g in the absence of oil and (d) filtration on Millipore filters.The intracellular content of water determined on cells from single rats weighing 120–150 g was 2.75 ± 0.55 μl/100 μl fat cells (± S.D., n = 30). The intracellular content of potassium, determined on cells from the same rats, was 252 ± 62 nmols/100 μl fat cells (± S.D., n = 30). The concentration of potassium in the intracellular water was calculated as 104 ± 15 mM (± S.D., n = 30).     

Determination of the kinetic constants of fructose transport and phosphorylation in the perfused rat liver

The kinetics of fructose uptake was determined in perfused rat liver during steady-state fructose elimination. On the basis of the corresponding values of fructose concentration in the affluent and in the effluent medium, and the fructose and ATP concentration in biopsies, the kinetics of membrane transport and intracellular phosphorylation in the intact organ was calculated according to a model system. Carrier-mediated fructose transport has a high $\text{K}{}_{\mathrm{<mtext>m</mtext>}}$ (67 mM) and $\text{V}$ (30 μmoles · min^?1 ·g^?1). The calculated kinetic constants of the intracellular phosphorylation were compared with values obtained with an acid-treated rat liver high speed supernatant (values given in parentheses). $\text{K}{}_{\mathrm{<mtext>m</mtext>}}$ with fructose 1.0 mM (0.7 mM), $\text{K}{}_{\mathrm{<mtext>m</mtext>}}$ with ATP 0.54 mM (0.37 mM), $\text{V}$ 10.3 μmoles · min^?1 · g^?1 (10.1 μmoles · min^?1 · g^?1, calculated on the basis of the highest measured rate of fructose uptake correcting the ATP concentration to saturating values). The kinetics of fructose uptake reveals that at Physiological fructose concentrations the membrane transport limits the rate of fructose uptake, thus protecting the liver from severe depletion of adenine nucleotides.     

Permeability of iodide in multilamellar liposomes modeled by two compartments and a reservoir

A previously published rate law for the diffusion of iodide from multilamellar egg phosphatidylcholine liposomes (Schullery, S.E. (1975) Chem. Phys. Lipids 49–58) is fitted to the relatively simple mathematical model of two compartments in series with a reservoir. All of the inner liposome compartments are assumed to behave as effectively one compartment in series with the liposome's outermost compartment. Based on this model, reasonable values are calculated for the fraction of the total solution trapped by liposomes which is in the outermost liposome compartment, 17%, and the permeability coefficient of iodide against isotonic, mixed iodide-chloride solution, $\text{2 · 10}{}^{\mathrm{?9}}\text{cm/s}$.     

Difference in microviscosity induced by different cholesterol levels in the surface membrane lipid layer of normal lymphocytes and malignant lymphoma cells

Microviscosity ($\text{}\text{?}$gh) in the surface membrane lipid layer of normal lymphocytes and malignant lymphoma cells, and in liposomes prepared from their lipid extracts, was determined with the aid of the fluorescence polarization properties of 1,6-diphenyl 1,3,5-hextriene embedded in it. The $\text{}\text{?}$gh values, both in intact cells and in the liposomes, are distinctively greater for normal lymphocytes than for the lymphoma cells, whereas the fusion activation energy in both types of cells and liposomes is 8 ± 0.5 kcal/mol. Determination of cholesterol revealed that its relative amount in a lymphoma cell is about half of that of a normal lymphocyte, a difference that may account for the above difference in fluidity. This thesis is supported by the observed changes in $\text{}\text{?}$gh, which follow artificial changes in cholesterol contents in the surface membrane of both cell types. Introduction of exogeneous cholesterol into the cell surface membranes was performed with lecithin-cholesterol (1:1) liposomes, and in lymphoma cells resulted in an increase of $\text{}\text{?}$gh to a level of normal lymphocytes. Extraction of native cholesterol from the cell surface membranes was carried out with lecithin liposomes, and in normal lymphocytes results in a decrease of $\text{}\text{?}$gh to a value similar to that of lymphoma cells. The induced changes in cholesterol contents are practically reversible for both cell types. By virtue of controlling the microviscosity of lipid layers, the level of cholesterol in cell surface membranes may play an important role in determining biological activities of normal and malignant cells.     

The effects of prostacyclin (PGI2) and 6-keto-PGF1α on bovine plasma progesterone and LH concentrations

Injections of 1 mg PGI₂ directly into the bovine corpus luteum significantly increased peripheral plasma progesterone concentrations within 5 min. Concentrations were higher in the PGI₂-treated heifers than in saline-injected controls between 5 and 150 min and at 3.5, 4, 5, and 7 h post-treatment. Levels tended to remain elevated through 14 h. Saline and 6-keto-PGF_1α were without effect on plasma progesterone levels. The luteotrophic effect of PGI₂ was not due to alterations in circulating LH concentrations. An $\text{in vitro}$ experiment assessed the effects of either PGI₂ alone or in combination with LH on progesterone production by dispersed luteal cells. Progesterone accumulation over 2 h for control, 5 ng LH, 1 μg PGI₂, 10 μg PGI₂, and 10 μg PGI₂ plus 5 ng LH averaged 99 ± 42, 353 ± 70, 152 ± 35, 252 ± 45, and 287 ± 66 ng/ml (n=4), respectively. Thus PGI₂ has luteotrophic effects on the bovine CL both $\text{in vivo}$ and $\text{in vitro}$.     

Inhibition by dexamethasone of the in vitro transport of 3-O-methylglucose into rat thymocytes

Reduced glucose transport across the plasma membrane and reduced phosphorylation may both be responsible for the early inhibitory effect of physiological concentrations of glucocorticoids on glucose uptake by rat thymocytes.The early inhibitory effects of glucocorticoids (5 · 10^?7 M dexamethasone) on glucose consumption and ¹⁴CO₂ formation from d-[U-¹⁴C]glucose were reproduced.The total uptake curve of 4.8 μM $\text{3-O-[}{}^{14}\text{C}\text{]}\text{methyl-}\text{d}\text{-glucose}$ was biexponential with $\text{t}\text{1}\text{2}$ of 1.1 min and 36 min, respectively, the rapid part comprising about 50% of the equilibrated intracellular water space. The latency of the effect of 5 · 10^?7 M dexamethasone on $\text{3-O-[}{}^{14}\text{C}\text{]}\text{methyl-}\text{d}\text{-glucose}$ uptake ranged from 15 to 100 min and the inhibition varied from 15 to 55% independently of the lag period. The effect of 3-O-methylglucose concentration on the initial uptake by steroid-responsive cell preparations was tested after 45 min of preincubation with or without 5 · 10^?7 M dexamethasone. In 12 experiments dexamethasone reduced V from 1.36 ± 0.16 mmol · min^?1 · l^?1 cell water to 0.81 ± 0.10 mmol · min^?1 · l^?1 cell water with insignificant change of K_m (6.0 mM versus 5.9 mM). Dexamethasone had similar effect after 90 or 120 min.The variabilities of control cell transport capacity, the lag period and the magnitude of the dexamethasone effect could not be accounted for by changes in pH, effects of cell density, concentrations of albumin, ethanol, nucleosides, pyruvate or correlated to age and sex of the rats. In conclusion the inhibition of glucocorticoids on glucose consumption by thymocytes appears to be an inhibited plasma membrane transport capacity.     

Kinetic parameters for the binding of p-nitrophenyl alpha-d-mannopyranoside to concanavalin A.

Binding of the chromogenic ligand p-nitrophenyl α-d-mannopyranoside to concanavalin A was studied in a stopped-flow spectrometer. Formation of the protein-ligand complex could be represented as a simple one-step process. No kinetic evidence could be obtained for a ligand-induced change in the conformation of concanavalin A, although the existence of such a conformational change was not excluded. The entire change in absorbance produced on ligand binding occurred in the monophasic process monitored in the stopped-flow spectrometer. The value of the apparent second-order rate constant (k_a) for complex formation (k_a = 54,000 s^?1m^? at 25 °C, pH 5.0, Γ/2 0.5) was independent of the protein concentration when the protein was in the range of 233–831 μm in combining sites and in excess of the ligand. The apparent first-order rate constant (k_?a) for dissociation of the complex was obtained from the rate constant for the decomposition of the complex upon the addition of excess methyl α-d-mannopyranoside (k_?a = 6.2 s^?1 at 25 °C, pH 5.0, Γ/2 0.5). The ratio $\text{k}{}_{\mathrm{<mtext>a</mtext>}}{}_{\mathrm{<mtext>?</mtext><mtext>a</mtext>}}$ (0.9 × 10₄m^?1) was in reasonable agreement with value of 1.1 ± 0.1 × 10⁴m^?1 determined for the equilibrium constant for complex formation by ultraviolet difference spectrometry. Plots of ln($\text{k}{}_{\mathrm{<mtext>a</mtext>}}\text{T}$) and ln($\text{k}{}_{\mathrm{<mtext>a</mtext>}}\text{T}$) vs $\text{1}\text{T}$ were linear (T is temperature) and were used to evaluate activation parameters. The enthalpies of activation for formation and dissociation of the complex are 9.5 ± 0.3 and 16.8 ± 0.2 kcal/mol, respectively. The unitary entropies of activation for formation and dissociation of the complex are 2.8 ± 1.1 and 1.3 ± 0.7 entropy units, respectively. These entropy changes are much less than those usually associated with substantial changes in the conformation of proteins.     

The relationship between 13, 14-dihydro-15-keto PGF2α and LH secretion in bulls

Half-life ($\text{t}\text{1}\text{2}$), volume of distribution (Vd)_and total body clearance (TBC) of 13, 14-dihydro-15-keto PGF_2α (PGFM) were measured in order to determine optimal sampling frequency for accurate measurement of PGFM. Three yearling Holstein bulls (349.2 ± 6.7 kg) and 3 yearling Holstein steers (346.7 ± 7.0 kg) were utilized in a 3 × 3 Latin square design. Animals were given 0, 25 or 50 μg PGF_2α I.V.; blood samples collected every 2 min and plasma PGFM determined. The $\text{t}\text{1}\text{2}$, Vd and TBC of PGFM were 2.3 ± .2 min, 43.3 ± 3.3 liters and 13.7 ± 1.9 liters/min, respectively and were similar for 25 and 50 μg doses. To determine the relationship between endogenous PGFM and LH secretion in bulls, blood samples were collected every 2 min for 12 h in 4 yearling Angus bulls (489.1 ± 11.6 kg). All animals elicited at least one LH surge and PGFM concentrations were measured in samples coincident with the LH surge. Mean plasma PGFM concentrations were greater prior to the LH surge than during the LH surge. In addition, mean plasma PGFM concentration and frequency of PGFM peaks appeared to increase prior to the LH surge suggesting an association between PGFM and pulsatile LH secretion in the bull.     

Interaction of bovine brain phospholipid exchange protein with liposomes of different lipid composition

The major phospholipid exchange protein from bovine brain catalyzes the transfer of phosphatidylinositol and phosphatidylcholine between rat liver microsomes and sonicated liposomes. The effect of liposomal lipid composition on the transfer of these phospholipids has been investigated. Standard liposomes contained phosphatidylcholine-phosphatidic acid (98:2, mol%); in general, phosphatidylcholine was substituted by various positively charged, negatively charged, or zwitterionic lipids. The transfer of phosphatidylinositol was essentially unaffected by the incorporation into liposomes of phosphatidic acid, phosphatidylserine, or phosphatidylglycerol (5–20 mol%) but strongly depressed by the incorporation of stearylamine (10–40 mol%). Marked stimulation (2–4-fold) of transfer activity was observed into liposomes containing phosphatidylethanolamine (2–40 mol%). The inclusion of sphingomyelin in the acceptor liposomes gave mixed results: stimulation at low levels (2–10 mol%) and inhibition at higher levels (up to 40 mol%). Cholesterol slightly diminished transfer activity at a liposome cholesterol/phospholipid molar ratio of 0.81. Similar effects were noted for the transfer to phosphatidylcholine from microsomes to these various liposomes. Compared to standard liposomes, the magnitude of $\text{K}{}_{\mathrm{<mtext>m</mtext>}}$ tended to increase for liposomes which depressed phospholipid transfer and to decrease for those which stimulated; little change was observed in the values of $\text{V}$. Single phospholipid liposomes of phosphatidylinositol were inhibitory when added to standard liposomes.     

Catecholamine-induced changes in transmembrane potential and temperature in normal and dystrophic hamster brown adipose tissue

Previous studies have shown that norepinephrine (NE) elicits trans-membrane potential changes in skeletal muscle cells from normal and dystrophic (BIO 14.6) hamsters, with the magnitude of these changes being significantly less in dystrophic cells. To determine if the decreased response of the dystrophic muscle cells reflects a more generalized phenomenon, the present study was designed to evaluate the effects of NE on membrane properties of brown adipocytes. $\text{In vivo}$ techniques using glass microelectrodes were similar to those used in the muscle studies. NE injection (2 to 5 μg/kg body wt, i.v.) into anesthetized hamsters was followed by membrane depolarization, the magnitude of which did not significantly differ in the dystrophic and normal adipocytes. For example, upon administration of 5 μg NE/kg body wt, the average depolarization was $\text{14.5 ± 1.3}\text{mV}\text{(}\text{X}\text{±}\text{S.E.}\text{)}$ for 20 dystrophic cells and 14.1 ± 1.8 mV for 18 normal cells. The depolarizations following i.v. infusion of isoproterenol and phenylephrine also had similar amplitudes in both normal and dystrophic cells. Despite this lack of difference in plasma membrane responses, NE induced a significantly smaller rise in interscapular brown fat temperature in the dystrophic (0.09°C) than in the normal hamsters (0.26°C) following administration of 5 μg NE/kg body wt. Thus, the decreased responsiveness to NE of dystrophic sarcolemma did not occur with the plasma membrane of brown adipocytes, although brown fat temperature changes in the dystrophic hamsters were decreased in amplitude.