Lymphocyte plasma membranes. III. Composition of lymphocyte plasma membranes from normal and immunized rats

Isolated plasma membranes of thymic and splenic lymphocytes from unimmunized and immunized rats of the inbred ACI and F344 strains were analyzed for chemical and enzymatic composition, for membrane protein patterns by polyacrylamide gel electrophoresis and for membrane-associated immunoglobulins. After immunization, the thymic and splenic lymphocyte membranes from F344 rat contained less carbohydrate and higher phospholipid contents than control animals. In both ACI and F344 inbred rat strains the membrane phospholipid to cholesterol weight ratio increased significantly after immunization. The electrophoretic patterns of solubilized membrane proteins and of iodinated external membrane proteins were similar in unimmunized and immunized animals.When thymic and splenic lymphocytes of normal or immunized animals were surface radioiodinated, solubilized in Triton X-100, NP-40 or 10 M urea in 1.5 M acetic acid and analyzed by immunoprecipitation, labeled IgM immunoglobulin was recovered from thymic lymphocytes but both labeled IgG and IgM were recovered from splenic lymphocytes. However, when unlabeled isolated plasma membranes were solubilized in 1% Triton X-100 and analyzed by immunodiffusion in agarose gels, both IgG and IgM were identified in thymic and splenic cells.     

Phosphoinositides of sheep platelets plasma membranes.

Phospholipid composition of sheep blood platelets and its various plasma membrane fractions have been analyzed. Based on their flotation rates in discontinuous sucrose density gradient centrifugation, three membrane fractions were isolated. 5'-Nucelotidase and alkaline phosphatase were distributed nearly equally in all the three membrane fractions. However these membrane fractions showed differences in the distribution of phosphatidyl ethanolamine, phosphatidyl choline and phosphoinositides. Phosphatidyl ethanolamine was predominant in fraction I (11.05 micrograms PLP/mg protein) while phosphatidyl choline was predominant in fractions II and III (110.10 and 68.30 micrograms PLP/mg protein respectively). Phosphatidyl inositol (Ptd-InsP) was equally distributed in all three membrane fractions. However, both Ptd-InsP and phosphatidyl inositol 4,5-bisphosphate were about 4-fold higher in fraction II (73.55 and 89.89 micrograms PLP/mg protein respectively).     

Lipids of bovine adrenal plasma membranes.

The lipid composition of a membrane fraction from bovine adrenal cortex was determined. This preparation has the capacity to bind adrenocorticotropic hormone and is enriched in adenylate cyclase and 5'-nucleotidase. The adrenal plasma membranes have a significantly higher lipid content (54.8%) than bovine liver plasma membranes and a surprisingly low proportion of this lipid is cholesterol (4.2%). The phospholipids comprise 76.4% of the total lipids and their composition if very similar to that of bovine liver membranes with the exception of sphingomyelin which comprises only 4.5% of the phospholipids in the adrenal preparation.     

Somatostatin binding to pituitary plasma membranes.

A method has been developed for the study of somatostatin binding to anterior pituitary plasma membranes. When 5×10^?9M [¹²⁵I]Tyr¹-somatostatin (SA 18 Ci/mmol) was incubated with isolated pituitary plasma membranes (protein = 100 μg), 13.6% of total radioactivity was bound excluding nonspecific binding. The Scatchard plot could be resolved into two distinct components and analyzed to yield: K₁diss = 3.3×10^?8M and K₂diss = 7.7×10^?6M. This binding was shown to be specific for somatostatin.     

Capsaicin effects on non-neuronal plasma membranes.

Capsaicin has been touted as a pharmacological tool specific for sensory afferent neurons and is widely used in neurophysiological studies. However, we have recently demonstrated that in concentrations commonly employed within the gastrointestinal tract, capsaicin inhibits platelet aggregation to at least three different stimuli. Since this was observed in a nerve free system it raised the question of how specific capsaicin is. In this communication we report that capsaicin has profound effects on physical properties of non-neuronal cell plasma membranes. These effects were observed while measuring the effect of capsaicin upon the fluidity of both intact cell membranes and a variety of purified membrane preparations. Membrane fluidity was assessed with the fluorescent probes diphenylhexatriene (DPH) and its trimethylamino derivative TMA-DPH and demonstrated concentration-dependent capsaicin effects. Furthermore, the effects were cell specific and for full expression required both intact cells and a non-lipid extractable component of the plasma membrane. These non-neuronal effects must be carefully considered when contemplating the explanation for capsaicin-induced effects.     

Characterization of rat liver cell plasma membranes.

A method is described by which bile canalicular membranes (BCM) can be prepared, together with canaliculus-free plasma membrane (PM), both essentially free of contamination. The recovery of both fractions together was estimated to be 46%. The concentrations of total lipid, total phospholipid and cholesterol were substantially greater in the BCM, and polyacrylamide gel electrophoresis revealed differences in protein composition. The differences in lipid and protein composition of these two plasma membrane fractions are presumably related to their very different physiological functions.     

Incorporation of ionic channels from yeast plasma membranes into black lipid membranes.

Recently, patch-clamping of yeast protoplasts has revealed the presence of plasma membrane K+ channels (Gustin, M. C., B. Martinac, Y. Saimi, M. R. Culberston, and C. Kung. 1986. Science (Wash. DC). 233:1195-1197). In this work we show that fusion of purified plasma membranes into planar bilayers allows the study of the yeast channels. The main cationic conductances detected were of 64 and 116 pS, however, larger and smaller conductances have been observed. The two main conductances were sensitive to the K+ channels blockers tetraethylammonium (TEA+) and Ba2+. Bionic experiments indicated that both conductances were K+ selective.