Abstract
Following our recent cloning of a novel γ-aminobutyric acid (GABA) receptor subunit geneResistance to dieldrin orRdl from the cyclodiene resistance locus inDrosophila melanogaster, we were interested in defining its pattern of expression during development. Here we report the raising of an anti-Rdl polyclonal antibody that recognizes a single protein of the expected 65 kDa size in immunoblots ofDrosophila head homogenates.In situ hybridization usingRdl cDNA probes and the anti-Rdl antibody shows thatRdl message and protein are highly expressed in the developing central nervous system (CNS) of 15–17 h embryos. Interestingly, despite the use of GABA in both the peripheral and CNS of insects,Rdl GABA receptor subunits appear to be confined to the CNS. Detailed immunocytochemistry ofDrosophila brain sections showed particularly strong anti-Rdl antibody staining in the optic lobes, ellipsoid body, fan shaped body, ventrolateral protocerebrum and the glomeruli of the antennal lobes. Results are compared with the distribution of staining observed in the insect CNS with antibodies against GABA itself and synaptotagmin, a synaptic vesicle protein.
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References
Ashburner, M. (1989) Drosophila:a laboratory manual. Cold Spring Harbor: Cold Spring Harbor Laboratory Press.
Buchner, E., Bader, R., Buchner, S., Cox, J., Emson, P. C., Flory, E., Heizmann, C. W., Hemm, S., Hofbauer, A. and Oertel, W. H. (1988) Cell-specific immuno-probes for the brain of normal and mutantDrosophila melanogaster.Cell Tissue Res.,253, 357–370.
Campos-Ortega, J. A. and Strausfeld, N. J. (1972) Columns and layers in the second synaptic region of the fly's visual system: the case for two superimposed neuronal architectures. InInformation processing in the visual systems of arthropods, ed. R. Wehner, pp. 31–36. Berlin: Springer-Verlag.
Campos-Ortega, J. A. and Hartenstein, V. (1985)The embryonic development of Drosophila melanogaster. Berlin: Springer-Verlag.
Chen, R., Belelli, D., Lambert, J. J., Peters, J. A., Reyes, A. and Lan, N. C. (1994) Cloning and functional expression of aDrosophila γ-aminobutyric acid receptor.Proc. Natl. Acad. Sci. USA,91, 6069–6073.
DiAntonio, A., Burgess, R. W., Chin, A. C., Deitcher, D. L., Scheller, R. H. and Schwarz, T. L. (1993) Identification and characterization ofDrosophila genes for synaptic vesicle proteins.J. Neurosci.,13, 4924–4935.
ffrench-Constant, R. H., Mortlock, D. P., Shaffer, C. D., MacIntyre, R. J. and Roush, R. T. (1991) Molecular cloning and transformation of cyclodiene resistance inDrosophila: an invertebrate GABAA receptor locus.Proc. Natl. Acad. Sci. USA,88, 7209–7213.
ffrench-Constant, R. H. and Rocheleau, T. (1993)Drosophila γ-aminobutyric acid receptor geneRdl shows extensive alternative splicing.J. Neurochem.,60, 2323–2326.
ffrench-Constant, R. H., Rocheleau, T. A., Steichen, J. C. and Chalmers, A. E. (1993) A point mutation in aDrosophila GABA receptor confers insecticide resistance.Nature,363, 449–451.
Henderson, J. E., Soderlund, D. M. and Knipple, D. C. (1993) Characterization of a putative γ-aminobutyric acid (GABA) receptor Β subunit gene fromDrosophila melanogaster.Biochem. Biophys. Res. Commun.,193, 474–482.
Homberg, U., Kingan, T. G. and Hildebrand, J. G. (1987) Immunocytochemistry of GABA in the brain and suboesophageal ganglion ofManduca sexta.Cell Tissue Res.,248, 1–24.
Hoskins, S. G., Homberg, U., Kingan, T. G., Christensen, T. A. and Hildebrand, J. G. (1986) Immunocytochemistry of GABA in the antennal lobes of the sphinx mothManduca sexta.Cell Tissue Res.,244, 243–252.
Kuffler, S. W. and Edwards, C. (1965) Mechanisms of gamma aminobutyric acid (GABA) action and its relation to synaptic inhibition.J. Neurophysiol.,21, 589–610.
Lee, H.-J., Rocheleau, T., Zhang, H.-G., Jackson, M. B. and ffrench-Constant, R. H. (1993) Expression of aDrosophila GABA receptor in a baculovirus insect cell system: functional expression of insecticide susceptible and resistant GABA receptors from the cyclodiene resistance geneRdl.FEBS Lett.,335, 315–318.
Matsumoto, S. G. and Hildebrand, J. G. (1981) Olfactory mechanisms in the mothManduca sexta: response characteristics and morphology of central neurons in the antennal lobes.Proc. R. Soc. Lond. B Biol. Sci.,213, 249–277.
Meyer, E. P., Matute, C., Streit, P. and NÄssel, D. R. (1986) Insect optic lobe neurons identifiable with monoclonal antibodies to GABA.Histochemistry,84, 207–216.
Olsen, R. W. and Tobin, A. J. (1990) Molecular biology of GABAA receptors.FASEB J.,4, 1469–1480.
Otsuka, M., Iversen, L. L., Hall, Z. W. and Kravitz, E. A. (1966) Release of gamma-aminobutyric acid from inhibitory nerves of lobster.Proc. Natl. Acad. Sci. USA,56, 1110–1115.
Restifo, L. L. and White, K. (1990) Molecular and genetic approaches to neurotransmitter and neuromodulator systems inDrosophila.Advances in Insect Physiology,22, 115–219.
Sattelle, D. B. (1990) GABA receptors of insects.Advances in Insect Physiology,22, 1–113.
Sattelle, D. B., Marshall, J., Lummis, S. C. R., Leech, C. A., Miller, K. W. P., Anthony, N. M. A., Bai, D., Wafford, K. A., Harrison, J. B., Chapaitis, L. A., Watson, M. K., Benner, E. A., Vassallo, J. G., Wong, J. F. H. and Rauh, J. J. (1992). γ-Aminobutyric acid and L-glutamate receptors of insect nervous tissue. InTransmitter Amino Acid Receptors, Structures, Transduction and Models for Drug Development, ed. E. A. Barnard and E. Losta, pp. 273–291. New York: Thieme Medical Publishers.
SchÄfer, S. and Bicker, G. (1986) Distribution of GABA-like immunoreactivity in the brain of the honeybee.J. Comp. Neurol.,246, 287–300.
Usherwood, P. N. R. and Grundfest, H. (1965) Peripheral inhibition in skeletal muscle of insects.J. Neurophysiol.,28, 497–518.
Watson, A. H. D. and Laurent, G. (1990) GABA-like immunoreactivity in a population of locust intersegmental interneurones and their inputs.J. Comp. Neurol.,302, 761–767.
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Aronstein, K., Ffrench-Constant, R. Immunocytochemistry of a novel GABA receptor subunitRdl inDrosophila melanogaster . Invertebrate Neuroscience 1, 25–31 (1995). https://doi.org/10.1007/BF02331829
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DOI: https://doi.org/10.1007/BF02331829