Search PubMed⌕ Search

Biomedical subjects

M M Poo

Publications and source records attributed to M M Poo.

At least 91 records · Page 5Linked to original sources
PubMed1984

Lateral electromigration and diffusion of Fc epsilon receptors on rat basophilic leukemia cells: effects of IgE binding.

We have used in situ electromigration and post-field relaxation (Poo, M.-m., 1981, Annu. Rev. Biophys. Bioeng., 10:245-276) to assess the effect of immunoglobulin E (IgE) binding on the lateral mobility of IgE-Fc receptors in the plasmalemma of rat basophilic leukemia (RBL) cells. Bound IgE sharply increased the receptor's electrokinetic mobility, whereas removal of cell surface neuraminic acids cut it to near zero. In contrast, we found only a small difference between the lateral diffusion coefficients (D) of vacant and IgE-occupied Fc receptors (D: 4 vs. 3 X 10(-10) cm2/s at 24 degrees C). This is true for monomeric rat IgE; with mouse IgE, the difference in apparent diffusion rates was slightly greater (D: 4.5 vs. 2.3 X 10(-10) cm2/s at 24 degrees C). This range of D values is close to that found in previous photobleaching studies of the IgE-Fc epsilon receptor complex in RBL cells and rat mast cells. Moreover, enzymatic depletion of cell coat components did not measurably alter the diffusion rate of IgE-occupied receptors. Thus, binding of fluorescent macromolecular probes to cell surface proteins need not severely impede lateral diffusion of the probed species. If the glycocalyx of RBL cells does limit lateral diffusion of the Fc epsilon receptor, it must act primarily on the receptor itself, rather than on receptor-bound IgE.

Animals↗
PubMed1984

Formation of electrical coupling between embryonic Xenopus muscle cells in culture.

Electrical coupling between embryonic Xenopus muscle cells in 1-5 day old cultures was studied after isolated cells were manipulated into contact for various periods. The coupling was examined by measuring the electrotonic spread of acetylcholine (ACh)-induced membrane depolarizations or of potential changes induced by intracellular current injection. In 1 day old culture, cells developed coupling rapidly after contact. Strong coupling was observed within 20 min after contact was made. The rate of coupling formation was age dependent. The percentage of cell pairs that established detectable coupling within 30 min of contact decreased from 66% in 1 day culture to 0% in 5 day culture. Older cells, when put into contact for prolonged periods, developed substantial coupling, suggesting that the age of the culture affects the rate of coupling formation rather than the final extent of coupling. Pre-treatment of older cells with colchicine, metabolic inhibitors, Ca2+ and Mg2+-free saline, or trypsin significantly increased the rate of coupling formation to a level close to that of younger cells. This suggests that the reduced rate of coupling was not due to a lack of membrane precursors for the intercellular channels, but was probably due to the appearance of extramembranous constraints for the channel assembly.

Animals↗
PubMed1984

Perturbation of the direction of neurite growth by pulsed and focal electric fields.

We have studied the orientation of neurite growth in the culture of embryonic Xenopus neurons in response to three types of extracellular electric fields: spatially uniform pulsed fields, focally applied steady (DC) fields, and focally applied pulsed fields. Under uniform pulsed fields, neurites showed a preferential orientation toward the cathode pole of the field in a manner similar to that previously found for DC fields. The extent of neurite orientation depended upon the duration, amplitude, and frequency of the pulse but appeared to be similar to that produced by a uniform DC field of an equivalent time-averaged field intensity. For square pulses of 5 msec duration, the minimal amplitude and frequency required to produce a detectable orientation of neurite growth over a period of 24 hr were 2.5 V/cm and 10 Hz, which correspond to a time-averaged field intensity of 125 mV/cm. Steady or pulsed focal fields were applied by passing a current through a micropipette placed near the growth cone of the neurite. Fields of negative polarity (current sink) were found to attract the growth cone, whereas fields of positive polarity (current source) were found to deflect the growth cone away from the pipette. The threshold DC current density needed at the growth cone to perturb its direction of growth within 15 min was 0.2 to 2 pA/micron2 (or 3 to 30 mV/cm); and for focal pulsed currents (pulse duration 5 msec), a typical combination of minimal pulse amplitude and frequency was 4 pA/micron2 and 10 Hz. This threshold focal current is similar to that which occurs at the synaptic cleft during active synaptic activity.

Animals↗
PubMed1983

Dissociated neurons from normal and mutant Drosophila larval central nervous system in cell culture.

A primary dissociated cell culture of Drosophila larval central nervous system is reported. Divisions of neuroblasts and vigorous outgrowth of neurites could be observed in culture. Within 24 hr cultured cells exhibited characteristic neuronal morphology and unimpaired ability to synthesize and accumulate acetylcholine. This cell culture system renders easy access to experimental analysis of normal neuronal properties and the altered mechanisms in neurological mutants. Single-channel currents induced by acetylcholine and regenerative action potentials were studied in the somata of the dissociated neurons. The appearance of Na channels in these cultured neurons was demonstrated by the cell lethality induced by veratridine and inhibition of the effect by tetrodotoxin. Dissociated neurons from a temperature-sensitive paralytic mutant napts, in which nerve conduction fails at high temperature, were studied in culture. Neuronal growth was not affected by this mutation, nor by tetrodotoxin. However, napts neurons showed greatly reduced sensitivity to veratridine even at 21 degrees C, a temperature at which napts individuals behave normally. This finding indicates expression of the napts phenotype at a level of isolated single cells and provides independent evidence that napts affects Na channel function.

Acetylcholine↗
PubMed1983

Rapid lateral diffusion of extrajunctional acetylcholine receptors in the developing muscle membrane of Xenopus tadpole.

We have studied the lateral diffusion of acetylcholine (ACh) receptors in the extrajunctional region of developing myotomal muscle cell membrane of Xenopus tadpoles by a technique of local inactivation. The myotomal muscle cell surfaces of Xenopus tadpoles were exposed to external solution by gently removing the skin of the tail. The density of ACh receptors was monitored by membrane depolarizations in response to iontophoretically applied pulses of ACh. A pulse of alpha-bungarotoxin was pressure ejected onto the exposed fiber surface, resulting in a rapid local inactivation of the ACh receptors. With time, the functional ACh receptors diffused into the region of inactivation, producing a recovery of ACh response. That the observed recovery of ACh sensitivity is due to diffusion of ACh receptors from the unexposed undersurface of the fiber to the inactivated region was evidenced by the following: (1) no recovery was observed following prolonged toxin application; (2) pretreatment of the muscle cells with concanavalin A, which cross-links and immobilizes ACh receptors, prevented recovery; (3) mapping of ACh response along the muscle cell axis showed that the recovery cannot be accounted for by diffusion along the longitudinal axis of the fiber; and (4) the diffusion coefficients observed after scaling the recovery rate with fiber radius fell within a small range (1.5 to 4.0 X 10(-9) cm2/sec), consistent with diffusion of ACh receptors around the fiber circumference. This finding of rapid lateral diffusion within developing tadpole myotomal muscle membrane supports the notion that the localization of ACh receptors induced by innervation could be achieved by a "diffusion-trap" mechanism where the nerve contact region serves as a trap for rapidly diffusing receptors in the membrane.

Acetylcholine↗
PubMed1982

Redistribution of cell surface receptors induced by cell-cell contact.

Cell surface lectin receptors underwent rapid redistribution after embryonic Xenopus myotomal muscle cells were manipulated into contact in culture. Soybean agglutinin (SBA) receptors became highly concentrated at the contact area and concanavalin A (Con A) and ricin receptors were depleted at the same region. The accumulation of SBA receptors was greatly reduced by the presence of SBA specific sugars in the incubating medium, by precontact binding of SBA to the surface and by lowering the temperature, but it was unaffected by prolonged treatments with metabolic inhibitors. It is culture-age dependent: older cultures showed a markedly reduced extent of accumulation, and the high accumulation resulting from contact made in younger cultures disappeared with time in culture. These findings are consistent with the notion that specific molecular interaction between the contacting surfaces results in a redistribution of preexisting rapidly diffusing surface receptors. In support of this notion, ligand-free SBA and Con A receptors were shown to be laterally mobile in the membrane, and at least a subpopulation of the SBA receptors contains physically distinct molecules from the Con A receptors. We suggest that such contact-induced redistribution of various surface components may play a role in the interaction between embryonic cells.

Animals↗
PubMed1982

Orientation of neurite growth by extracellular electric fields.

Extracellularly applied steady electric fields of 0.1 to 10 V/cm were found to have marked effects on the neurite growth of single dissociated Xenopus neurons in culture: (1) neurites facing the cathode showed accelerated growth, while the growth of those facing the anode was reduced. Neurites growing relatively perpendicular to the field axis were prompted to curve toward the cathode. (2) More neurites appeared to be initiate from the cathodal side of the cell. (3) The number of neurite-bearing neurons per culture and the average neurite length were increased. These effects are absent in cultures treated with electric fields of similar strength but alternating polarity and cannot be attributed either to a gradient of extracellular diffusible substances or to the flow of culture medium produced by the field. The field effects are reversible: (1) removal of the electric field resulted in the loss of neurite orientation in a few hours and (2) reversal of the polarity of the electric field led to a rapid reversal in the neurite orientation. To determine the cellular loci of these field effects, we treated the neurons with a number of pharmacological agents or altered their ionic environments. Incubation with concanavalin A (Con A) was found to abolish these filed effects completely. Since the binding of Con A to the neuronal surface was shown to prevent field-induced accumulation of the Con A receptors toward the cathodal side of these neurons, our finding is accumulation of the Con A receptors toward the cathodal side of these neurons, our finding is consistent with the notion that cathodal accumulation of growth-controlling surface glycoproteins by the field is the underlying mechanism of the field-induced orientation of neurite growth toward the cathode.

Animals↗
PubMed1981

Localization of cell membrane components by surface diffusion into a "trap".

Randomly distributed cell membrane components may become localized toward a specific region of the surface as a results of cell-cell contact or the cell's exposure to extracellular ligands. The mechanism for this localization process is unknown. In the present study, we investigated the plausibility of a passive mechanisms, namely that a local region of the cell surface serves as a "trap" for diffusing membrane proteins. Based on a model of spherical cell with a single circular trap on the surface, we derived the equation describing the surface density distribution and the average lifetime of the trappable molecules in the trap-free region of the membrane. This surface-trap theory was then used to analyze our experimental finding on the rapid localization of muscle surface soybean agglutinin receptors induced by cell-cell contact in culture. THe result indicates that the rate of localization of these receptors toward the cell-cell contact site can be accounted for by assuming that the receptors possess a diffusion coefficient of about 2.5 X10(-9) cm2/s (range: 1.2-9.3X10(-9) cm2/s) before they are trapped at the contact site. Independent measurement of the rate of lateral diffusion of these receptors yielded a lateral diffusion coefficient of about 1.9 X 10(-9) cm2/s (range 1.2-2.7 X10(-9) cm2/s), a value within the range of that predicted by the rate of localization. We thus conclude that lateral diffusion of mobile membrane components toward a local trap is a plausible mechanism for their localization induced by local surface modulation.

Animals↗
PubMed1981

The role of electro-osmosis in the electric-field-induced movement of charged macromolecules on the surfaces of cells.

The surfaces of most cells bear a net negative charge. The imposition of an electric field parallel to the surface of the cell should produce, therefore, an electro-osmotic flow of fluid towards the cathodal side of the cell. Our analysis of a simple model of the cell surface indicates that a negatively charged mobile macromolecule will be swept by this electro-osmotic flow of fluid to the cathodal side of the cell if its zeta potential, zeta 1, is less negative than the zeta potential of the cell surface, zeta 2. Conversely, if zeta 2 is less negative than zeta 1, the negatively charged macromolecule will accumulate at the anodal side of the cell. Our experimental results demonstrate that concanavalin A (Con A) receptors on embryonic muscle cells normally accumulate at the cathodal side of the cell, but that they can be induced to accumulate at the anodal side of the cell by preincubating the myotubes either with neuraminidase, a treatment that removes negatively charged sialic acid residues, or with the lipid diI, a treatment that adds positive charges to the surface of the cell. Addition of the negatively charged lipid monosialoganglioside (GM1), on the other hand, enhances the accumulation of Con A receptors at the cathodal side of the cell.

Animals↗
PubMed1979

Neurites grow faster towards the cathode than the anode in a steady field.

We explanted fragments of embryonic chick dorsal root ganglia on to polylysine coated glass and cultured them in a medium containing one unit of nerve growth factor plus enough methylcellulose to give viscosities from 0.01-3,000 poise. We allowed them to grow out in the absence of a field, and then selected explants with halos of neurites which were relatively dense, relatively symmetrical, and practically free of glial cells. These selected explants were then exposed to electrical fields of up to 140 mV/mm for some hours. In media with viscosities of one poise or less, the field some times dragged the central cell mass of an explant towards the anode. However, in cases where the central cell mass did not move, fields of 70-140 mV/mm induced that sector of each neurite halo which faced the cathode to grow out several times faster than the one facing the anode.

Animals↗
PubMed1978

Lateral electrophoresis and diffusion of Concanavalin A receptors in the membrane of embryonic muscle cell.

A uniform electric field of 10 V/cm applied across the surface of embryonic toad Xenopus muscle cells results in the asymmetric accumulation of concanavalin A (Con A) receptors toward one side of the cells within 10 min, as visualized by postfield fluorescent Con A labeling. This field produces an extracellular voltage difference of 20 mV across these 20-microns wide cells. The effect is reversible in two respects: (a) Additional exposure of the cell to the same field of opposite polarity for 10 min completely reverses the asymmetric accumulation to the other side of the cell. (b) Relaxation occurs after the removal of the field and results in complete recovery of the uniform distribution in 30 min. Both the accumulation and the recovery movements are independent of cell metabolism, and appear to be electrophoretic and diffusional in nature. The threshold field required to induce a detectable accumulation by the present method is between 1.0 and 1.5 V/cm (corresponding to a voltage difference of 2-3 mV across a 20-microns wide cell). The electrophoretic mobility of the most mobile population of nonliganded Con A receptors is estimated to be about 2 x 10(-3) microns/s per V/cm, while their diffusion coefficient is in the range of 4-7 x 10(-10) cm2/s. Extensive accumulation of the Con A receptors by an electric field results in the formation of immobile aggregates. The Con A receptors appear to consist of a heterogeneous population of membrane components different in their charge properties, mobility, and capability in forming aggregates.

Animals↗
PubMed1977

Relations between ameboid movement and membrane-controlled electrical currents.

We have studied the pattern of electrical currents through amebas (mainly Chaos chaos) with an ultrasensitive extracellular vibrating probe. Amebas drive both steady currents and current pulses through themselves. Relatively steady current with an average surface density of 0.1-0.2 muA/cm2 enters the rear quarter of an ameba and leaves its pseudopods. Streaming reversals are preceded by changes in this current pattern and the region with the largest new inward current becomes the new tail. Ion substitution studies suggest that some of the steady inward current is carried by calcium ions. Characteristic stimulated pulses of current sometimes follow the close approach of the vibrating probe to the side of an advancing pseudopod. Such a pulse enters the cytoplasm through a small patch of membrane near the probe (and seems to leave through the adjacent membrane), is usually followed by hyaline cap and then by pseudopod initiation, is calcium dependent, lasts about 5-10 s, and has a peak density of about 0.4 muA/cm2. Spontaneous pulses of similar shape and duration may enter or leave any part of an animal. They are much less localized, tend to have higher peak densities, and occur in physiological salt solutions at about 0.2-4 times per minute. Retraction of a pseudopod is always accompanied or preceded by a spontaneous pulse which leaves its sides.

Amoeba↗
PubMed1977

Passive signal propagation and membrane properties in median photoreceptors of the giant barnacle.

1. The light-induced electrical responses of barnacle photoreceptors spread decrementally along the cells' axons. The decay of the depolarizing and hyperpolarizing components of the visual signal was studied by recording intracellularly from single receptor axons of the median ocellus of the giant barnacle.2. The resistance of the photoreceptor neurone decreases markedly when the cell is depolarized with respect to its dark resting potential of -60 mV. This rectification results in differential attenuation of the depolarizing and hyperpolarizing components of the visual signal as they spread down the axon. Consequently, the visual signal entering the synaptic region is conspicuously distorted.3. Bathing the photoreceptor axons in sodium-free or calcium-free saline or in isotonic sucrose does not significantly affect the spread of the visual signal to the terminals. Thus the signal is not amplified by an ionic mechanism along the axon.4. Membrane characteristics of the photoreceptor for hyperpolarizing voltage changes were estimated from (a) the ratio of the amplitudes of the visual signals recorded simultaneously in the axon and in the soma, (b) the time constant, and (c) the input resistance of the cell. All three independent measurements are consistent with a length constant 1 to 2 times the total length of the cell (lambda = 10-18 mm) and an unusually high membrane resistivity of about 300 kOmega cm(2). This resistivity enables the receptor potential to spread passively to the terminal region.5. Electron microscopic examination of receptor axons reveals an investment of glial lamellae, but demonstrates neither unusual structures which would lead to a high apparent membrane resistivity, nor junctions between cells which would seal off the extracellular space. Thus the observed high resistivity appears to be an intrinsic property of the receptor membrane.

Animals↗
PubMedDate not supplied

Topographical rearrangement of acetylcholine receptors alters channel kinetics.

Plasma membranes are dynamic structures of proteins and lipids. Protein-protein or protein-lipid interactions within the membrane are believed to have important roles in many membrane functions, including ion transport, enzyme activity and signal reception. The acetylcholine (ACh) receptor-channel complex in skeletal muscle membrane is one of the best known integral membrane proteins. Its ion transport function is accessible to direct measurement at the single-channel level by the use of the 'giga-seal' patch recording technique. Here we used an in situ electrophoresis technique to rearrange the topography of pre-existing ACh receptor-channels in the muscle membrane, and measured the single-channel kinetics of ACh-activated channels in two different molecular environments within the membrane: those in the diffusely distributed region and those in the ACh receptor clusters induced by the applied field. We found that the channel kinetics are significantly prolonged in the ACh receptor cluster compared with the non-clustered region of the same cell. This result strongly supports the notion that the function of a membrane ionic channel depends on the local molecular environment.

Animals↗