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Improved stability of Drosophila larval neuromuscular preparations in haemolymph-like physiological solutions

Journal of Comparative Physiology A volume 175, pages 179–191 (1994)Cite this article

Abstract

Neuromuscular preparations from third instar larvae of Drosophila are not well-maintained in commonly used physiological solutions: vacuoles form in the muscle fibers, and membrane potential declines. These problems may result from the Na∶K ratio and total divalent cation content of these physiological solutions being quite different from those of haemolymph. Accordingly haemolymph-like solutions, based upon ion measurements of major cations, were developed and tested. Haemolymph-like solutions maintained the membrane potential at a relatively constant level, and prolonged the physiological life of the preparations. Synaptic transmission was well-maintained in haemolymph-like solutions, but the excitatory synaptic potentials had a slower time course and summated more effectively with repetitive stimulation, than in standard Drosophila solutions. Voltage-clamp experiments suggest that these effects are linked to more pronounced activation of muscle fiber membrane conductances in standard solutions, rather than to differences in passive muscle membrane properties or changes in postsynaptic receptor channel kinetics. Calcium dependence of transmitter release was steep in both standard and haemolymph-like solutions, but higher external calcium concentrations were required for a given level of release in haemolymph-like solutions. Thus, haemolymph-like solutions allow for prolonged, stable recording of synaptic transmission.

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Abbreviations

HL:

haemolymph-like

References

  • Atwood HL, Govind CK, Wu C-F (1993) Neuromuscular junction ultrastructure of ventral abdominal muscles in Drosophila larvae. J Neurobiol 24: 1008–1024

    Google Scholar 

  • Begg M, Cruickshank WJ (1963) A partial analysis of Drosophila larval haemolymph. Proc R Soc Edinburgh 68B: 215–236

    Google Scholar 

  • Broadie KS, Bate M (1993) Development of larval muscle properties in the embryonic myotubes of Drosophila melanogaster. J Neurosci 13: 167–180

    Google Scholar 

  • Croghan PC, Lockwood APM (1960) The composition of the haemolymph of the larva of Drosophila melanogaster. J Exp Biol 37: 339–343

    Google Scholar 

  • Crossley CA (1978) The morphology and development of the Drosophila muscular system. In: Ashburner M, Wright TRF (eds) The genetics and biology of Drosophila, vol. 2b. Academic Press, New York, pp 449–559

    Google Scholar 

  • Delgado R, Latorre R, Labarca P (1992) K+-channel blockers restore synaptic plasticity in the neuromuscular junction of dunce, a Drosophila learning and memory mutant. Proc R Soc Lond B 250: 181–185

    Google Scholar 

  • Dodge FA, Rahamimmoff R (1967) Cooperative action of calcium ions in transmitter release at the neuromuscular junction. J Physiol (Lond) 193: 419–432

    Google Scholar 

  • Dudel J (1981) The effect of reduced calcium on the quantal unit current and release at the crayfish neuromuscular junction. Pflügers Arch 391: 35–40

    Google Scholar 

  • Elkins T, Ganetzky B, Wu C-F (1986) A Drosophila mutation that eliminates a calcium-dependent potassium current. Proc Natl Acad Sci USA 83: 8415–8419

    Google Scholar 

  • Florkin M, Jeuniaux C (1974) Hemolymph: Composition. In: Rockstein M (ed) The physiology of insecta, Vol V. Academic Press, New York, pp 255–308

    Google Scholar 

  • Gho M, Ganetzky B (1992) Analysis of repolarization of presynaptic motor terminals in Drosophila larvae using potassiumchannel blocking drugs and mutations. J Exp Biol 170: 93–111

    Google Scholar 

  • Haugland FN, Wu C-F (1990) A voltage-clamp analysis of gene dosage effects of the Shaker locus on larval muscle potassium currents in Drosophila. J Neurosci 10: 1357–1371

    Google Scholar 

  • Hille B (1992) Ionic channels of excitable membranes, 2 ed., Sinauer Associates Inc, Sunderland MA

    Google Scholar 

  • Ikeda K, Kaplan WD (1970) Patterned neural activity of a mutant of Drosophila melanogaster. Proc Natl Acad Sci USA 66: 765–772

    Google Scholar 

  • Irving SN, Miller TA (1980) Ionic differences in ‘fast’ and ‘slow’ neuromuscular transmission in body wall muscles of Musca domestica larvae. J Comp Physiol 135: 291–298

    Google Scholar 

  • Irving SN, Wilson RG, Osborne MP (1979) Studies on L-glutamte in insect haemolymph III. Amino acid analyses of the haemolymph of various arthropods. Physiol Entomol 4: 231–240

    Google Scholar 

  • Jan LY, Jan YN (1976) Properties of the larval neuromuscular junction in Drosophila melanogaster. J Physiol (Lond) 262: 189–214

    Google Scholar 

  • Jan LY, Jan YN, Dennis MJ (1977) Two mutations of synaptic transmission in Drosophila. Proc R Soc Lond B 198: 189–214

    Google Scholar 

  • Johansen J, Halpern ME, Johansen KM, Keshishian H (1989) Stereotypie morphology of glutamatergic synapses on identified muscle cells of Drosophila larvae. J Neurosci 9: 710–725

    Google Scholar 

  • Keshishian H, Chiba A, Chang TN, Halfon MS, Harkins EW, Jarecki J, Wang L, Anderson M, Cash S, Halpern ME, Johansen J (1993) Cellular mechanisms governing synaptic development in Drosophila melanogaster. J Neurobiol 24: 757–787

    Google Scholar 

  • Mallart A (1993) Calcium-dependent modulation of the facilitation of transmitter release at neuromuscular junctions of Drosophila. J Physiol (Paris) 87: 83–88

    Google Scholar 

  • Mallart A, Angaut-Petit D, Bourret-Poulain C, Ferrus A (1991) Nerve terminal excitability and neuromuscular transmission in T(X:Y)V7 and Shaker mutants of Drosophila melanogaster. J Neurogenet 7: 75–84

    Google Scholar 

  • Meyran JC, Sellem E, Graf F (1993) Variations in haemolymph ionized calcium concentrations in the crustacean Orchestia cavimana during the moult cycle. J Insect Physiol 39: 413–417

    Google Scholar 

  • Piek T, Njio KD, Mantel P (1973) Effects of anions and dinitrophenol on resting membrane potentials of insect muscle fibers. J Insect Physiol 19: 2373–2392

    Google Scholar 

  • Piek T, Mantel P, Wijsman JPM (1977) Effects of monocarboxylates and external pH on some parameters of insect muscle fibres. J Insect Physiol 23: 773–777

    Google Scholar 

  • Rheuben MB (1972) The resting potential of moth muscle fibre. J Physiol (Lond) 25: 1–25

    Google Scholar 

  • Seabrook GR, Duce IR, Irving SN (1989) Spontaneous and evoked quantal neurotransmitter release at the neuromuscular junction of the larval housefly, Musca domestica. Pflügers Arch 414: 44–51

    Google Scholar 

  • Sigaard-Andersen O, Thode J, Fogh-Andersen N (1983) What is “ionized calcium”? Scand J Clin Invest 43: 11–16

    Google Scholar 

  • Singh S, Wu C-F (1989) Complete separation of four potassium currents in Drosophila. Neuron 2: 1325–1329

    Google Scholar 

  • Singh S, Wu C-F (1990) Properties of potassium currents and their role in membrane excitability in Drosophila larval muscle fibers. J Exp Biol 152: 59–76

    Google Scholar 

  • Sink H, Whitington PM (1991) Location and connectivity of abdominal motoneurons in the embryo and larva of Drosophila melanogaster. J Neurobiol. 22: 298–311

    Google Scholar 

  • Suzuki N, Kano M (1977) Development of action potential in larval muscle fibers in Drosophila melanogaster. J Cell Physiol 93: 383–388

    Google Scholar 

  • Van der Meer JM, Jaffe LF (1983) Elemental composition of the perivitelline fluid in early Drosophila embryos. Dev Biol 95: 249–252

    Google Scholar 

  • Wareham AC, Duncan CJ, Bowler K (1973) Bicarbonate ions and the resting potential of cockroach muscle: Implications for the development of suitable saline media. Comp Biochem Physiol 45A: 239–246

    Google Scholar 

  • Wareham AC, Duncan CJ, Bowler K (1974) The resting potential of cockroach muscle membrane. Comp Biochem Physiol 48A: 765–797

    Google Scholar 

  • Wareham AC, Duncan CJ, Bowler K (1975) The resting potential of the muscle membrane of moths. Comp Biochem Physiol 52A: 295–298

    Google Scholar 

  • Wu C-F, Ganetzky B (1980) Genetic alteration of nerve membrane excitability in temperature-sensitive paralytic mutants of Drosophila melanogaster. Nature 286: 814–816

    Google Scholar 

  • Wu C-F, Ganetzky B (1988) Genetic and pharmacological analysis of ion channels in Drosophila. In: Lunt GG (ed) Neurotox 88: Molecular basis of drug and pesticide action. Elsevier Science Publishers B.V, Amsterdam, pp 311–323

    Google Scholar 

  • Wu C-F, Ganetzky B, Jan LY, Jan YN, Benzer S (1978) A Drosophila mutant with a temperature-sensitve block in nerve conduction. Proc Natl Acad Sci USA 75: 4047–4051

    Google Scholar 

  • Yamaoka K, Ikeda K (1988) Electrogenic responses elicited by transmembrane depolarizing current in aerated body wall muscles of Drosophila melanogaster larvae. J Comp Physiol A 163: 705–714

    Google Scholar 

  • Zhong Y, Wu C-F (1991) Altered synaptic plasticity in Drosophila memory mutants with a defective cyclic AMP cascade. Science 251: 198–201

    Google Scholar 

  • Zhong Y, Wu C-F (1993) Differential modulation of potassium currents by cAMP and its long-term and short-term effects: dunce and rutabaga mutants of Drosophila. J Neurogenet 9: 15–27

    Google Scholar 

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Authors and Affiliations

  1. Department of Physiology, University of Toronto, M5S 1A8, Toronto, Ontario, Canada

    B. A. Stewart & H. L. Atwood

  2. Department of Biological Sciences, University of Iowa, 52242, Iowa City, IA, USA

    H. L. Atwood, J. J. Renger, J. Wang & C.-F. Wu

Authors
  1. B. A. Stewart
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  2. H. L. Atwood
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  3. J. J. Renger
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  4. J. Wang
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  5. C.-F. Wu
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Stewart, B.A., Atwood, H.L., Renger, J.J. et al. Improved stability of Drosophila larval neuromuscular preparations in haemolymph-like physiological solutions. J Comp Physiol A 175, 179–191 (1994). https://doi.org/10.1007/BF00215114

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  • DOI: https://doi.org/10.1007/BF00215114

Key words

  • Drosophila
  • Neuromuscular
  • Haemolymph
  • Membrane potential
  • Synaptic potential