ABSTRACT
Na(+)-K(+)-ATPase is localized to the basolateral cell surface of most epithelial cells. Conflicting results regarding the intracellular trafficking of Na(+)-K(+)-ATPase in Madin-Darby canine kidney cells have been reported, with delivery to both apical and basolateral membranes or exclusively to the basolateral cell surface. We examined the delivery and steady-state distribution of Na(+)-K(+)-ATPase in the amphibian epithelial cell line A6 using an antibody raised against Na(+)-K(+)-ATPase alpha-subunit and sulfo-N-hydroxysuccinimidobiotin to tag cell surface proteins. The steady-state distribution of the Na(+)-K(+)-ATPase was basolateral, as confirmed by immunocytochemistry. Delivery of newly synthesized Na(+)-K(+)-ATPase to the cell surface was examined using [35S]methionine and [35S]cysteine in a pulse-chase protocol. After a 20-min pulse, the alpha-subunit and core glycosylated beta-subunit were present at both apical and basolateral cell surfaces. The alpha-subunit and core glycosylated beta-subunit delivered to the apical cell surface were degraded within 2 h. Mature alpha/beta-heterodimer was found almost exclusively at the basolateral surface after a 1- to 24-h chase. These data suggest that immature Na(+)-K(+)-ATPase alpha-subunit and core glycosylated beta-subunits are not retained in the endoplasmic reticulum of A6 cells and apparently lack sorting signals. Mature Na(+)-K(+)-ATPase is targeted to the basolateral surface, suggesting that basolateral targeting of the protein is conformation dependent.
Subject(s)
Cell Membrane/enzymology , Protein Processing, Post-Translational , Sodium-Potassium-Exchanging ATPase/metabolism , Animals , Cell Line , Cell Membrane/ultrastructure , Dogs , Epithelial Cells , Epithelium/enzymology , Kidney/enzymology , Models, Biological , Sodium-Potassium-Exchanging ATPase/biosynthesis , Sodium-Potassium-Exchanging ATPase/isolation & purification , Xenopus laevisABSTRACT
Cross-correlation analysis of interdependence of the background spike activity was carried out for pairs of adjacent neurons simultaneously recorded in the incubated slices of the neocortex of guinea-pig. Statistical correlation of spike discharges was detected in 16 out of 26 recorded pairs of the neurons. Significant correlation was observed mainly in the range of +/- 100 ms from the null point. Cross-correlation had symmetric or asymmetric maxima up to 150 ms long and negative shifts up to 200 ms long. More complex positive-negative types of cross-correlations were also obtained. The data were compared to those known from other authors for the intact brain. The contribution of intrinsic intracortical interactions and extrinsic afferent influences in these correlations of activity is discussed.
Subject(s)
Motor Cortex/physiology , Neurons/physiology , Animals , Guinea Pigs , In Vitro Techniques , Membrane Potentials/drug effects , Membrane Potentials/physiology , Motor Cortex/drug effects , Neurons/drug effects , Sodium Glutamate/pharmacology , Statistics as Topic , Time FactorsABSTRACT
Visual cortex slices of guinea pig were maintained in vitro. Background neuronal activity was recorded extracellularly. Five types of the activity were revealed: single regular, single irregular, bursting, group and mixed discharges. The regular pacemaker-type discharges were encountered more often than other types. Such regular activity was never observed in the neocortex in vivo. The type of the background activity of one and the same neuron may change with time.
Subject(s)
Neurons/physiology , Tissue Survival , Visual Cortex/physiology , Animals , Culture Techniques , Electrophysiology , Guinea Pigs , Time FactorsABSTRACT
Background neuronal discharges were recorded extracellularly in guinea-pig cerebral cortex slices maintained in vitro. Six types of activity were classified: 1--regular single discharges (38.5%), 2--irregular spikes (6.5%), 3--bursts (6.5%), 4--burst-single discharge (mixed) activity (26.5%), 5--group discharges (2.5%), 6--multineuronal volleys (18.5%). Characteristic distributions of interspike intervals and autocorrelograms corresponded to each of these types of activity. Regular pacemaker-type discharges were most peculiar under conditions of small volumes of the grey matter and lack of afferentation. Such regular activity was never observed in normal or isolated neocortex.
Subject(s)
Motor Cortex/physiology , Somatosensory Cortex/physiology , Action Potentials , Animals , Guinea Pigs , In Vitro TechniquesABSTRACT
Antiserum to neurotoxin-2 from the venom of the cobra was obtained by immunization of rabbits via the injection of the toxin into the lymphatic nodes and by two re-immunizations via the injection of the toxin into the blood and intramuscularly. Using scores of micrograms of the toxin, specific antisera with high antibody titer were obtained. In spite of high specificity of separate stages of the reactions, using indirect immune fluorescence technique, no specific fluorescence was found in the end-plates of the diaphragm, in which "intact" cholinoreceptors were bound to neurotoxin-2. It is suggested that antigenic determinant of the toxin is involved into the formation of a firm link with the receptors, this process resulting in "masking" the determinant and in loss of its ability to react with antibodies.
