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B N Christensen

Publications and source records attributed to B N Christensen.

35 records · Page 2Linked to original sources
PubMed1986

Voltage-clamp analysis of currents produced by glutamate and some glutamate analogues on horizontal cells isolated from the catfish retina.

Horizontal cells isolated from the catfish retina were exposed to radiolabeled glutamate, glycine, gamma-aminobutyric acid (GABA), and sucrose to determine if the enzymatic dissociation procedure altered the high-affinity uptake mechanism for GABA and generally reduced membrane selectivity. As in the intact retina, isolated cells could transport GABA but not the other substances. The horizontal cells were voltage clamped using a single low-resistance patch-type electrode. The acidic amino acid L-glutamate, and its analogues kainate and quisqualate, were applied to the cell by pressure ejection from a nearby pipette. All three agonists produced inward currents that reversed near O mV. Quisqualate produced a current with a similar time course as glutamate, but the time course of the response to kainate was faster. The agonists N-methyl-D-aspartate and L-aspartate had little effect on the membrane conductance. The current-to-voltage (I-V) relationship for all three agonists was nonlinear when the membrane potential was hyperpolarized. The nonlinearity was, at least in part, a result of the decreased response to the three agonists. Removal of Mg did not alter this nonlinear relationship. When the inward potassium rectifier was blocked with 100 microM Ba, the response to glutamate was increased compared with the control experiment before block by barium; however, the I-V relationship was still highly nonlinear. Thus glutamate block of the inward potassium current cannot account entirely for the nonlinear I-V. The increase in membrane permeability to specific ions in the presence of an agonist was determined by ion substitution experiments and measuring the shift in the reversal potential. The three agonists appear to increase the membrane permeability to cations but not to anions. The amino acid antagonists cis-2,3-piperidine dicarboxylic acid (PDA) and D-glutamyl glycine (DGG) were bath applied to test their ability to block the depolarizing effects of glutamate. DGG had no measureable effect at 100 microM concentration, whereas PDA reversibly reduced the glutamate response at 1 mM concentration although block was incomplete. Isolated horizontal cells responded to bath-applied glutamate in concentrations of 10-500 microM. In concentrations of glutamate greater than 50 microM, when the membrane potential was held at the resting potential, the inward current reached a maximum followed by a decrease to a steady-state level. This apparent time-dependent desensitization at high agonist concentrations was at least partially removed when Mg was removed from the bathing solution.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1986

Excitable properties and voltage-sensitive ion conductances of horizontal cells isolated from catfish (Ictalurus punctatus) retina.

External horizontal cells were enzymatically dissociated from intact catfish (Ictalurus punctatus) retina and pipetted onto a small chamber attached to the stage of an inverted phase-contrast microscope. Individual horizontal cells were recognized by their large size and restricted dendritic arborization. Low-resistance (3-12 M omega) patch-type electrodes were used to record intracellular potentials and to pass current across the cell membrane under either current or voltage-clamp conditions. The average resting potential of isolated horizontal cells was -67 V + 6.9 mV (mean +/- SD, n = 40). At the resting potential, the cell membrane appears to be mainly permeable to K. A depolarizing current step evoked an action potential in the cell. The maximum rate of rise of the action potential (dV/dt) in normal physiological solution was 6.5 +/- 1.8 V/s (means +/- SD, n = 24) and was reduced to 1.2 +/- 0.39 V/s (means +/- SD, n = 9) in 1-10 micron tetrodotoxin (TTX) and 3.2 +/- 1.4 V/s (means +/- SD, n = 6) in Ca-free solution. The maximum dV/dt was reduced in 10 mM extracellular K concentration [K]o to about half of that seen in standard saline, and values in 30 or 80 mM [K]o were similar to that measured in TTX. Following an action potential, the membrane potential reached a plateau potential of + 17.4 +/- 8.1 mV (means +/- SD, n = 17) and remained depolarized for variable periods of time lasting from less than a second to a few minutes. When the plateau potential was long lasting, the cell repolarized slowly and upon reaching zero rapidly repolarized to the original resting potential. The duration of the plateau potential decreased or was absent in saline containing one of the following calcium channel antagonists: La, Cd, Co, or Ni. The voltage-clamp technique was used to identify the membrane currents responsible for the membrane potential changes seen under current clamp. Experiments were carried out using either a single or two individual electrodes. Fast and steady-state inward currents were recorded from isolated horizontal cells in the voltage range between -20 and +20 mV. These currents were a result of increased membrane conductance to both Na and Ca ions. The Na channels are inactivated at depolarized potentials and are TTX sensitive. Ca channels are partially inactivated at depolarized potentials. The Ca conductance is decreased by Cd, Co, Ni, and La. Ba can substitute for Ca in the channel.(ABSTRACT TRUNCATED AT 400 WORDS)

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PubMed1985

White noise analysis of cable properties of neuroblastoma cells and lamprey central neurons.

The integrative properties of two kinds of excitable cells, a cultured neuroblastoma cell and the lamprey giant interneuron, are described using a white-noise impedance method. The impedance functions were fitted with a neuron model consisting of an isopotential soma plus a single equivalent dendritic process, which contained up to 16 equal elements. The frequency-domain characteristics of both the passive and active conductances were used to estimate the dendritic-to-soma areas, the electrotonic length of an equivalent dendrite, the membrane time constant, and the relaxation time constants associated with the voltage-dependent conductances. The effect of differing degrees of synaptic input was simulated by localizing the synaptically activated conductances to the soma, a point at the end of the dendrite, or the entire dendritic membrane.

Animals↗
PubMed1983

Sodium and calcium currents measured in isolated catfish horizontal cells under voltage clamp.

A low resistance suction microelectrode was used to record intracellularly from enzymatically dissociated horizontal cells obtained from catfish retina (Ictalurus punctatus). This microelectrode was connected to a voltage clamp circuit and the transmembrane currents were recorded during depolarizing and hyperpolarizing clamp pulses. Two fast transient inward currents were recorded, one sensitive to tetrodotoxin and one insensitive to this drug. A slower and persistent tetrodotoxin insensitive inward current was also recorded. The results indicate that separate membrane conductances exist for both sodium and calcium ions. We suggest that the action potential recorded from isolated horizontal cells is generated by both sodium and calcium currents.

Animals↗
PubMed1983

Distribution of electrotonic synapses on identified lamprey neurons: a comparison of a model prediction with an electron microscopic analysis.

1. Intracellular recordings of the monosynaptic excitatory postsynaptic potential (EPSP) were made from an identified giant interneuron in the lamprey spinal cord following stimulation of a single identified presynaptic giant axon. The EPSP had both electrotonic and chemical components. 2. The giant interneuron and sometimes the giant axon were labeled with horseradish peroxidase (HRP). Using combined light and electron microscopy (EM), gap junctions formed by the axon on the HRP-labeled dendritic trees of seven interneurons were identified and their position marked on dendritic diagrams made from tracings of the cells. 3. The total number of gap junctions on a giant interneuron varied from a minimum of 9 to a maximum of 23. The number of gap junctions on a branch of a dendritic tree varied from 1 to 9. For all seven neurons the closest gap junction was located approximately 65 microns from the soma and the furthest 280 microns from the soma. 4. When gap junction position was plotted as a function of electrotonic distance from the soma, the variability in their distribution decreased significantly. Synaptic location was found to be on the average about 0.5 space constants from the soma, and this value compared favorably with that determined from an analysis of half-width and rise time of the electrotonic synaptic potential for these neurons. 5. These results suggest that the presynaptic axon tends to make, on the average, synaptic contact at the same electrical distance from the soma.

Animals↗
PubMed1981

Direct electrical connections between transient amacrine cells in the catfish retina.

Transient amacrine cells were identified by their intracellularly recorded response to flashes of light. These cells typically respond with a transient depolarization, often followed by a steady-state response during the stimulus. When two electrodes were placed in different transient amacrine cells, current of either polarity passed through one electrode produced a steady-state voltage change that was recorded by the electrode in the nearby cell. Following identification of the physiological type, transient amacrine cells were injected with horseradish peroxidase and the tissue was processed for light and electron microscopy. Both conventional chemical synaptic junctions and gap junctions were found to connect amacrine cells.

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PubMed1980

A quantitative analysis of ultrastructural changes induced by electrical stimulation of identified spinal cord axons in the larval lamprey.

Microelectrodes filled with horseradish peroxidase (HRP) were used to label single identified giant axons in the isolated lamprey spinal cord. Subsequent to the iontophoretic injection of HRP, the spinal cord was stimulated at repetition rates of 20-30/s and the activity in labelled axons monitored. Immediately following failure of the action potential, the spinal cord was fixed by immersion and processed for light and electron microscopy. Electron micrographs were taken of synaptic contacts made by the labelled axons. Several quantitative measures were made from each synapse using a digitizing tablet interfaced with a digital computer. These measures included vesicle number (VN), vesicle area (VA), length of differentiated membrane (DM), vesicle density (VD = VN/VA), vesicle frequency (VF = VN/DM), and a relative measure of the amount of vesicle membrane added to the axolemma during the stimulation period, the curvature ratio (CR). Measures from 106 stimulated synapses were compared with 134 synapses from injected but unstimulated giant axons. The results from these experiments suggest that measurable ultrastructural changes occur during transmitter release at identified C.N.S. synapses, which are consistent with the hypothesis of synaptic vesicle recycling.

Action Potentials↗
PubMed1979

The relationship between some measures of synaptic ultrastructure as a function of distance from the soma on lamprey reticulospinal neurons.

Synaptic junctions located on the dendrites of lamprey (Petromyzon marinus) reticulospinal neurons labelled with intracellularly-injected horseradish peroxidase were studied. The normal ultrastructure of the synaptic junctions was defined and several quantitative measures made from each junction in order to test the hypothesis that distally-located synapses are ultrastructurally different from those located at proximal dendritic sites. A total of 820 contacts from one neuron and 279 from a second neuron ranging from 20 to 340 microns from the soma were quantified. The vast majority of the presynaptic endings contained round, clear-cored vesicles and formed an asymmetrical membrane differentiation with the postsynaptic dendrite. A small fraction of the population contained flattened or pleomorphic vesicles and these synapses were equally distributed with respect to distance from the soma. Many of the terminals contained a few large dark- and clear-cored vesicles. Four quantitative measures of each synaptic contact were made. These included vesicle number, length of differentiated membrane, vesicle area and terminal area. Four ratios relating the different quantitative measures were also calculated. Each ratio or measurement from the synaptic junctions was plotted as a function of distance from the soma to determine if differences existed at any distance. It was found that synaptic junctions are uniformly similar and that distal junctions did not differ significantly (P greater than 0.05) from those at proximal dendritic sites. It is concluded that if distal synapses do compensate for their remote location they do this is some other way, possibly by increasing the number of synaptic contacts made by each presynaptic axon.

Animals↗
PubMed1979

Estimates of cable parameters in lamprey spinal cord neurones.

1. Two micro-electrodes were used to penetrate giant interneurones in the isolated lamprey spinal cord. A brief (50--100 microsec) current pulse was applied to one electrode while the other recorded the voltage transient response. 2. A formal analysis of the voltage transient was achieved by the simplifying reduction of each neurone. Somas were treated as a parallel combination of resistance and capacitance. Dendrite trees were reduced to an equivalent cylinder (Rall, 1959). 3. The voltage transients were analysed according to the procedure suggested by Jack & Redman (1971b) to estimate the cable parameters governing the passive propagation of transmembrane potentials. Membrane time constant (tau m), dendritic to soma conductance ratio (rho 00), and electrotonic length (L) of the equivalent cylinder were estimated from these data. 4. In thirty-two interneurones it was possible to determine the membrane time constant, but rho 00 and L were determined in only twenty-two. 5. For the twenty-two neurones in which all cable parameters were estimated, the electrotonic length of the equivalent cylinder was similar to that found for cat spinal motoneurones (1--2 space constants). 6. Simulations of the voltage transient using the Rall model of the motoneurone as developed by Jack & Redman (1971b) resulted in a voltage response which closely ditted the experimental data. 7. These results suggest that the Rall model of the motoneurone accurately describes the propagation of passive transmembrane potentials in lamprey spinal cord neurones. It is further concluded that the time constant for soma and dendritic membrane is similar in these neurones.

Animals↗
PubMed1979

Localization of synaptic input on dendrites of a lamprey spinal cord neurone from physiological measurements of membrane properties.

1. Composite excitatory post-synaptic potentials (e.p.s.p.s) resulting from electrotonic and chemical synaptic junctions were recorded from eighteen interneurones following stimulation of the I2 burster axon in the isolated lamprey spinal cord. 2. In each cell, the half-width of the electrotonic e.p.s.p. was measured and used, together with the cable parameters estimated for the same neurone, to locate the position of synaptic contact made by the I2 axon on the dendrites of the interneurone. The synaptic location ranged from 0.05 to 1.35 space constants with a mean of 0.46. 3. The synaptic potential was simulated using the Rall model of the neurone. When compared with the experimentally recorded e.p.s.p. with the same half-width, the rise-time of the simulated synaptic potential was found to be faster. By changing the value of synaptic distance and/or synaptic current duration the half-width, rise-time, and decay of the simulated synaptic potential fit closely the experimental e.p.s.p. The range of synaptic distance estimated from the simulation decreased considerably (0.2--0.7 space constants; mean 0.52). 4. Direct comparison of synaptic location estimated from histological tracings of dendritic trees from these same cells injected with horseradish peroxidase compared favourably with synaptic location estimated from the simulations. 5. These results support the hypothesis that functionally similar presynaptic axons make synaptic connexions at the same electrotonic distance from the soma on functionally similar post-synaptic cells. This occurs in the face of large variations in physical distance for these same synaptic contacts.

Animals↗
PubMed1976

Morphological correlates of synaptic transmission in lamprey spinal cord.

The dye Procion brown was used to identify in the light and electron microscope, synaptic contacts made between monosynaptically coupled neurons in the lamprey spinal cord whose synaptic interaction had been recorded. Synaptic contacts were made on different dendrites of the postsynaptic cell at different distances from the soma. Some of the contacts were made on dentritic spines and some on the smooth shaft of the dentrites. Serial sections through synaptic contacts made on dendritic processess of the postsynaptic cells were used for three-dimensional reconstruction of the synapses using computer graphics techniques. The computer reconstructions and detailed examination of the serial EM micrographs revealed the large proliferation of membrane involved in making these en passant synapses as well as the morphological changes due to stimulation of the presynaptic axon. These changes include depletion of synaptic vesicles and formation of complex vesicles and synaptic cisternae. Besides chemical synaptic contacts, four electrotonic contacts were located, confirming the mixed electrochemical synaptic response recorded from the postsynaptic cell. The mean quantum content was estimated and compared with the estimate of the available transmitter pool, assuming the quantal release hypothesis applies at these synapses. The total transmitter pool was estimated by counting all synaptic vesicles in all synaptic contacts. It was estimated that about 6% of the total transmitter pool is available for release at these synapses. This compares with less than 1% at the neuromuscular junction and about 20% at sympathetic synapses. These results support the hypothesis that synaptic vesicles may be recycled as described by Heuser and Reese (22) at the neuromuscular junction. Ongoing studies are investigating the effect on a variety of synaptic junctions to stimulation for different periods of time of presynaptic axons. The methods described in this study can also be used to test the models of synaptic interaction on dendritic trees described by Rall (39) and Jack and Redman (24).

Action Potentials↗
PubMed1973

Procion brown: an intracellular dye for light and electron microscopy.

Procion browvn has been used to inject lamprey giant interneutros. The dye is opaque to both light and electrons, and imparts a pink color to the neuron in situ during iontophoresis. It has the advantage over cobaltolus chloride that postinjection treatment with ammonium sulfide is eliminated and the integrity of the surrounding neuropil is ensured.

Animals↗
PubMed1970

Estimates of probability of transmitter release at the mammalian neuromuscular junction.

1. Intracellular records of end-plate potentials (e.p.p.s) were obtained from curarized neuromuscular junctions in the rat diaphragm.2. Statistical estimates of the quantal release probability (p) were made at individual junctions from measurements of the means and variances of e.p.p. amplitudes at two different levels of Ca concentration. It was assumed that the release process was binomial and that the number of quanta available for release (n) was independent of external Ca.3. These estimates of p were compared with those obtained by measuring depression of e.p.p. amplitudes after single conditioning shocks and assuming that the depression was due to depletion of n. The statistical estimates were consistently smaller.4. This disparity, plus the additional observation that depression was not linearly related to the number of quanta released by one or more conditioning shocks, suggested that depression was not due entirely to depletion and that the depletion assumption led to an over-estimate of p.5. A third method of estimating p from measurements of the decline of e.p.p. amplitudes during rapid stimulation also appeared to result in an over-estimate.

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