Worm Breeder's Gazette 13(5): 32 (February 1, 1995)
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
Institute of Neuroscience, 1254 University of Oregon, Eugene, OR, 97403. Using simple modifications of existing techniques, I have recently made intracellular recordings from neurons and muscles in C. elegans. Larvae (approx. L2) were glued with cyanoacrylate adhesive to a coverslip coated with a moist agarose film.1 The coverslip formed the bottom of the recording chamber, which was filled with a physiological saline and viewed on an inverted microscope using Nomarski optics. Animals were dissected in a two-step procedure. First, internal pressure was relieved by nicking the cuticle in the mid-gut using a tungsten needle. Second, either the pharynx or a small bouquet of neurons was exposed by making a nick in the cuticle of the head. Intracellular microelectrodes were used to record from the pharynx and patch electrodes were used to record from neurons. Microelectrode recordings from muscles of the spontaneously pumping pharynx revealed rhythmic depolarizations with an amplitude of 50 to 70 mV and a duration of several hundred milliseconds. These events closely resembled action potentials previously recorded from the pharynx of Ascaris,2 indicating that important aspects of physiological function are retained after gluing and dissection. As a first step in understanding the basic operating principles of the C. elegans nervous system, I have concentrated primarily on whole-cell voltage-clamp recordings. Twenty-one whole-cell or perforated patch recordings have been made so far. Neuronal input capacitance ranged from 0.1 to 2.0 pF. The low end of this range is the capacitance expected of an isolated L2 soma,3 while the upper end is the capacitance expected of an L2 neuron with a process about 50 mm long. The apparent neuronal input resistance ranged from 0.1 to 7.1 G ohms. Patch clamp methods systematically underestimate capacitance and resistance in small neurons. Nevertheless, these data indicate the membrane time constant, which determines how fast a neuron responds to its inputs, is at least 14 ms. They also suggest the axonal space constant, which determines how far an input signal propagates passively, is at least 150 mm. This means that interneurons confined to the nerve ring should be effectively isopotential. Two classes of neurons could be distinguished by differences in their voltage-dependent currents. Cells of the first class had sustained outward currents but no inward currents. Cells of the second class had a transient outward current and also a small, sustained inward current. Because the inward current activates more rapidly and at lower clamp voltages than the outward current, these cells may be capable of regenerative potentials. Until now, all recordings have been from unidentified neurons. I am hopeful, however, that by using GFP4 labeled worms I will be able to record from identified neurons. If so, it should be possible to determine whether different types of neurons have different electrophysiological properties and to correlate these differences with the behavioral roles predicted by anatomical and laser ablation experiments. Moreover, by crossing GFP animals with mutant strains, I hope to record from identified cells in mutants. Thus, it should be possible to combine the electrophysiological and the genetic analysis of behavior at the cellular level in individual neurons. 1. Avery, L., Raizen, D., and Lockery, S.R. (1995) Electrophysiological Methods. In Epstein, H.F. and Shakes, D.C. (eds.) C. Elegans: Modern Biological Analysis of an Organism. Academic Press, Orlando (in press). 2. Byerly L., Masuda, M.O. (1979). J. Physiol. 288:263-284. 3. David Hall, unpublished data. 4. Chalfie, M., Tu, Y., Euskirchen, G., Ward, W.W. and Prasher, D.C. (1994). Science 263:802-5.