Cable properties of a straight neurite of a leech neuron probed by a voltage-sensitive dye.

Abstract

We measured a time-resolved map of electrical activity in a thin straight neurite (1. (24). The 100 outputs were fed into current-voltage converters (OPA 121; Burr-Brown, Filderstadt, F.R.G., with 100-Ma feedback resistors, time constant 0.4 ms) and eventually into differential sample-and-hold amplifiers. Ten diodes were selected, and their signals were stored on a tape recorder (Racal, Bergisch Gladbach, F.R.G.).The protocol of a measurement was as follows: (i) Adjustment of a segment of the neurite along a row of 10 diodes, (ii) record baselines in the dark, (iii) record fluorescence with open shutter (after a 5-ms interval) and then switch to the sample-and-hold mode with the total fluorescence as a reference. (iv) Record and store fluorescence with electrical stimulation for 70 ms: A positive current (2-4 nA) was injected for 25 ms to elicit an action potential (amplitude 60-75 mV). After an interval of 10 ms, a negative Gaussian voltage (amplitude around -100 to -160 mV) was induced by a negative and positive current pulse. (v) After an interval of 5 s the fluorescence without stimulation was recorded for 70 ms and stored (baseline of bleaching). To overcome the limited range of the diode array (-85 pum) we displaced the neuron along a row of diodes several times-starting with the terminal segment-and repeated cycles (i-v). A complete set of data was sampled from the tape at an interval of0.1 ms and a resolution of 12 bit and read into a microcomputer. The fluorescence changes without stimulation were fitted by an exponential function and subtracted from the changes with stimulation; the results were divided by the total fluorescences. The transients of depolarization were corrected for the decreased amplitude of the action potential (from 75 to 60 mV) during a set of measurements with displaced neurites.Then all transients of depolarization were normalized to constant amplitude. The transients of hyperpolarization were scaled by the same factors. Finally the signals were smoothed Abbreviation: N cell, nociceptive neuron. 4604The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

Effect of Neuritic Cables on Conductance Estimates for Remote Electrical Synapses

“…We found an average cytoplasmic resistivity R of about 400 ⍀cm, a value far higher than the classical 35 ⍀cm for the squid giant axon (Hodgkin and Huxley 1952) but similar to values reported for other narrower neurites such as the 250 ⍀cm assigned to cultured leech neurons (Fromherz and Müller 1994;Fromherz and Vetter 1992) and the 340 and 300 ⍀cm found for rat hippocampal neurons (Major et al 1994;Meyer et al 1997). The average neuritic membrane conductance G ϭ 0.035 mS/cm 2 that led to the best fits in three experiments is similar to values found with imaging techniques in cultured leech neurons (G ϭ 0.05 mS/cm 2 ) (Fromherz and Müller 1994) and cultured rat hippocampal pyramidal neurons (G ϭ 0.07 mS/ cm 2 ) (Meyer et al 1997).…”

“…In cells whose somatic membrane properties differ from those of the neurites (Durand 1984, Kawato 1984, this parameter identification problem has, however, been demonstrated to be ill-posed in that very small errors in measured data can lead to large and unpredictable errors in parameter estimates (White et al 1992). To avoid that problem, we took advantage of the simple geometry and good optical accessibility of cultured Lymnaea nerve cells and determined their cable properties from voltage-sensitive dye measurements of voltage transients in the entire cell rather than at the cell body alone (Fromherz and Müller 1994). We estimated the electrical parameters by fitting voltage transients simulated with a passive multi-compartment model to voltage transients reconstructed from fluorescence measurements throughout the cell and transients recorded directly at the cell body (Fig.…”

“…For dye excitation, the light from a Xenon lamp was passed through a 450 to 190 nm band-pass onto the cell. The fluorescent light emitted by the dye in the somatic and neuritic membrane was long-pass filtered at 560 nm and recorded at a spatial resolution of 4 -5 m as previously described (Fromherz and Müller 1994;Meyer et al 1997) while hyperpolarizing current pulses were injected into the cell.…”

“…The technique allows the design of pairs of neurons that share a single point of contact with an electrical synapse at the terminal of two straight neurites-a geometry that corresponds to the one considered in our theoretical approach. Isolating the neurons from their surrounding tissue and restricting their neuritic tree to a straight cable has the additional advantage of facilitating the computation of the neuritic cable properties from high-resolution optical-imaging data obtained with voltage-sensitive dyes (Fromherz and Müller 1994). This alternative way of obtaining cable properties uses voltage transients observed throughout the neuritic cable rather than transients measured at the cell body alone.…”

Optical Recording of Action Potentials with Second-Harmonic Generation Microscopy

“…The current capabilities demonstrated here are most immediately applicable to reproducibly stimulated systems, as are often used for current optical V m recording techniques (Zecevic, 1996). It should be feasible to investigate spike initiation zones (Zecevic, 1996) and AP propagation properties (Fromherz and Muller, 1994;Meyer et al, 1997) deep in intact ganglia, where one-photon methods are not appropriate. Repeated line scans at varying spatial positions over processes will allow for the generation of high-resolution time series movies of AP propagation, leading to the experimental analysis of electrical propagation properties at complex structures, such as axon bifurcations and varicosities.…”