Immunoreactivity against choline acetyltransferase, γ‐aminobutyric acid, histamine, octopamine, and serotonin in the larval chemosensory system of Dosophila melanogaster

We have studied the distribution of choline acetyltransferase (ChAT), γ‐aminobutyric acid (GABA), histamine, octopamine and serotonin in the larval chemosensory system of Drosophila melanogaster. Colocalization at the confocal level with green fluorescent protein (GFP) or Tau‐GFP reporters, expressed in selected P[GAL4] enhancer trap lines, was used to identify the cells making up these neurotransmitters. As in the adult fly, larval olfactory afferents project into the (larval) antennal lobe (LAL), where they synapse onto local interneurons and projection neurons, whereas gustatory afferents terminate essentially in the tritocerebral‐subesophageal (TR‐SOG) region. We demonstrate that the neuropils of the LAL and the TR‐SOG are immunoreactive to ChAT and GABA. In addition, serotonin‐ and octopamine‐immunoreactive fibers are present in the LAL. ChAT immunostaining is localized in subsets of olfactory and gustatory afferents and in many of the projection neurons. In contrast, GABA is expressed in most, and perhaps all, of the local interneurons. Serotonin immunoreactivity in the LAL derives from a single neuron that is situated close to the LAL and projects to additional neuropil regions. Taken together, these findings resemble the situation in the adult fly. Hence, given the highly reduced numbers of odorant receptor neurons in the larva, as shown in a previous study (Python and Stocker [2002] J. Comp. Neurol. 445:374–387), the larval system may become an attractive model system for studying the roles of neurotransmitters in olfactory processing. J. Comp. Neurol. 453:157–167, 2002. © 2002 Wiley‐Liss, Inc.

[1]  D. Nässel.  Functional roles of neuropeptides in the insect central nervous system , 2000, Naturwissenschaften.

[2]  P. Distler,et al.  Olfactory Bulb and Antennal Lobe , 1990 .

[3]  R. Stocker,et al.  Adult‐like complexity of the larval antennal lobe of D. melanogaster despite markedly low numbers of odorant receptor neurons , 2002, The Journal of comparative neurology.

[4]  John R. Carlson,et al.  A Novel Family of Divergent Seven-Transmembrane Proteins Candidate Odorant Receptors in Drosophila , 1999, Neuron.

[5]  J. Hildebrand,et al.  Distribution of acetylcholinesterase activity in the deutocerebrum of the sphinx moth Manduca sexta , 1995, Cell and Tissue Research.

[6]  J. Hildebrand,et al.  Olfactory control of behavior in moths: central processing of odor information and the functional significance of olfactory glomeruli , 2004, Journal of Comparative Physiology A.

[7]  A. Brand,et al.  In vivo dynamics of axon pathfinding in the Drosophilia CNS: a time-lapse study of an identified motorneuron. , 1998, Journal of neurobiology.

[8]  U. Homberg,et al.  Neuroactive Substances in the Antennal Lobe , 1999 .

[9]  L. Luo,et al.  Representation of the Glomerular Olfactory Map in the Drosophila Brain , 2002, Cell.

[10]  M. Hammer.  An identified neuron mediates the unconditioned stimulus in associative olfactory learning in honeybees , 1993, Nature.

[11]  P. Salvaterra,et al.  Localization of choline acetyltransferase‐expressing neurons in Drosophila nervous system , 1999, Microscopy research and technique.

[12]  A. Baumann,et al.  Molecular and pharmacological properties of insect biogenic amine receptors: lessons from Drosophila melanogaster and Apis mellifera. , 2001, Archives of insect biochemistry and physiology.

[13]  I. Meinertzhagen,et al.  Synaptic organization of the mushroom body calyx in Drosophila melanogaster , 2002, The Journal of comparative neurology.

[14]  R. Stocker,et al.  A central neural circuit for experience-independent olfactory and courtship behavior in Drosophila melanogaster , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[15]  S. Kreissl,et al.  Octopamine‐like immunoreactivity in the brain and subesophageal ganglion of the honeybee , 1994, The Journal of comparative neurology.

[16]  Y. Kidokoro,et al.  Octopamine inhibits synaptic transmission at the larval neuromuscular junction in Drosophila melanogaster , 1999, Brain Research.

[17]  N. Perrimon,et al.  Targeted gene expression as a means of altering cell fates and generating dominant phenotypes. , 1993, Development.

[18]  R. Greenspan,et al.  Natural behavior polymorphism due to a cGMP-dependent protein kinase of Drosophila. , 1997, Science.

[19]  G. Boulianne,et al.  Green fluorescent protein as a vital marker and reporter of gene expression in Drosophila. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[20]  J. Hildebrand,et al.  A novel serotonin-immunoreactive neuron in the antennal lobe of the sphinx moth Manduca sexta persists throughout postembryonic life. , 1987, Journal of neurobiology.

[21]  G. Bicker,et al.  Distribution of GABA‐like immunoreactivity in the brain of the honeybee , 1986, The Journal of comparative neurology.

[22]  G. Laurent,et al.  Impaired odour discrimination on desynchronization of odour-encoding neural assemblies , 1997, Nature.

[23]  M. Monastirioti,et al.  Biogenic amine systems in the fruit fly Drosophila melanogaster , 1999, Microscopy research and technique.

[24]  D. Malun.  Synaptic relationships between GABA-immunoreactive neurons and an identified uniglomerular projection neuron in the antennal lobe of Periplaneta americana: a double-labeling electron microscopic study , 2004, Histochemistry.

[25]  M. Sokolowski,et al.  The foraging gene affects adult but not larval olfactory-related behavior in Drosophila melanogaster , 1998, Behavioural Brain Research.

[26]  Liqun Luo,et al.  Target neuron prespecification in the olfactory map of Drosophila , 2001, Nature.

[27]  P. Hiesinger,et al.  Three‐dimensional reconstruction of the antennal lobe in Drosophila melanogaster , 1999, The Journal of comparative neurology.

[28]  Richard Axel,et al.  An Olfactory Sensory Map in the Fly Brain , 2000, Cell.

[29]  J. Hildebrand,et al.  Immunocytochemistry of GABA in the antennal lobes of the sphinx moth Manduca sexta , 2004, Cell and Tissue Research.

[30]  M. Cobb.  What and how do maggots smell? , 1999 .

[31]  E. Buchner,et al.  Choline acetyltransferase-like immunoreactivity in the brain of Drosophila melanogaster , 1986, Cell and Tissue Research.

[32]  K. White,et al.  Development of serotonin-containing neurons in Drosophila mutants unable to synthesize serotonin , 1986, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[33]  Gerd Bicker.  Sources and targets of nitric oxide signalling in insect nervous systems , 2001, Cell and Tissue Research.

[34]  U. Homberg.  Distribution of Neurotransmitters in the Insect Brain , 1994 .

[35]  J. Hildebrand,et al.  Insect Olfaction , 1999, Springer Berlin Heidelberg.

[36]  E. Buchner,et al.  Histamine is a major mechanosensory neurotransmitter candidate in Drosophila melanogaster , 1993, Cell and Tissue Research.

[37]  E. Buchner,et al.  Autoradiographic localization of [3H]choline uptake in the brain of Drosophila melanogaster , 1983, Neuroscience Letters.

[38]  G. Bicker,et al.  Histochemistry of acetylcholinesterase and immunocytochemistry of an acetylcholine receptor‐like antigen in the brain of the honeybee , 1989, The Journal of comparative neurology.

[39]  U Homberg,et al.  Immunocytochemistry of GABA in the central complex of the locust Schistocerca gregaria: Identification of immunoreactive neurons and colocalization with neuropeptides , 1999, The Journal of comparative neurology.

[40]  R. Stocker,et al.  Neuroblast ablation in Drosophila P[GAL4] lines reveals origins of olfactory interneurons. , 1997, Journal of neurobiology.

[41]  K. White,et al.  Serotonin‐containing neurons in Drosophila melanogaster: Development and distribution , 1988, The Journal of comparative neurology.

[42]  A. Hofbauer,et al.  Histamine-like immunoreactivity in the visual system and brain of Drosophila melanogaster , 1991, Cell and Tissue Research.

[43]  Linda B. Buck,et al.  Information coding in the olfactory system: Evidence for a stereotyped and highly organized epitope map in the olfactory bulb , 1994, Cell.

[44]  U. Müller.  THE NITRIC OXIDE SYSTEM IN INSECTS , 1997, Progress in Neurobiology.

[45]  P. Guerin,et al.  Neurophysiological and behavioural evidence for an olfactory function for the dorsal organ and a gustatory one for the terminal organ in Drosophila melanogaster larvae. , 2000, Journal of insect physiology.

[46]  D. Nässel,et al.  Aminergic neurons in the brain of blowflies and Drosophila: dopamine- and tyrosine hydroxylase-immunoreactive neurons and their relationship with putative histaminergic neurons , 2004, Cell and Tissue Research.

[47]  Gerd Bicker.  Histochemistry of classical neurotransmitters in antennal lobes and mushroom bodies of the honeybee , 1999, Microscopy research and technique.

[48]  P. Salvaterra,et al.  Differential regulation of choline acetyltransferase expression in adult Drosophila melanogaster brain. , 1996, Journal of neurobiology.

[49]  J. Hirsh,et al.  Temporal and spatial development of serotonin and dopamine neurons in the Drosophila CNS. , 1994, Developmental biology.

[50]  P. Taghert.  FMRFamide neuropeptides and neuropeptide‐associated enzymes in Drosophila , 1999, Microscopy research and technique.

[51]  Richard Axel,et al.  Spatial Representation of the Glomerular Map in the Drosophila Protocerebrum , 2002, Cell.

[52]  Andrey Rzhetsky,et al.  A Spatial Map of Olfactory Receptor Expression in the Drosophila Antenna , 1999, Cell.

[53]  M. Monastirioti,et al.  Octopamine immunoreactivity in the fruit fly Drosophila melanogaster , 1995, The Journal of comparative neurology.

[54]  D. Sattelle,et al.  Localization in the Nervous System of Drosophila Melanogaster of a C‐Terminus Anti‐Peptide Antibody to a Cloned Drosophila Muscarinic Acetylcholine Receptor , 1995, Journal of neuroendocrinology.

[55]  L. Tolbert,et al.  Development of an identified serotonergic neuron in the antennal lobe of the moth and effects of reduction in serotonin during construction of olfactory glomeruli. , 1995, Journal of neurobiology.

[56]  D. Schild,et al.  Chemosensory Information Processing , 1990, NATO ASI Series.

[57]  T. Kitamoto,et al.  Immunocytochemical study of choline acetyltransferase in Drosophila melanogaster: An analysis of cis‐regulatory regions controlling expression in the brain of cDNA‐transformed flies , 1995, The Journal of comparative neurology.

[58]  R. N. Singh,et al.  Fine structure of the sensory organs of Drosophila melanogaster Meigen larva (Diptera : Drosophilidae) , 1984 .

[59]  R. Stocker.  Drosophila as a focus in olfactory research: Mapping of olfactory sensilla by fine structure, odor specificity, odorant receptor expression, and central connectivity , 2001, Microscopy research and technique.

[60]  J. Hildebrand,et al.  Structure and function of the deutocerebrum in insects. , 1989, Annual review of entomology.

[61]  Andrey Rzhetsky,et al.  A Chemosensory Gene Family Encoding Candidate Gustatory and Olfactory Receptors in Drosophila , 2001, Cell.

[62]  D. Nässel,et al.  Histaminelike immunoreactive neurons innervating putative neurohaemal areas and central neuropil in the thoraco‐abdominal ganglia of the flies Drosophila and Calliphora , 1990, The Journal of comparative neurology.

[63]  T. Kitamoto,et al.  Regulation of choline acetyltransferase/lacZ fusion gene expression in putative cholinergic neurons of Drosophila melanogaster. , 1995, Journal of neurobiology.

[64]  D. Nässel.  Histamine in the brain of insects: a review , 1999, Microscopy research and technique.

[65]  P. Salvaterra,et al.  Analysis of choline acetyltransferase protein in temperature sensitive mutant flies using newly generated monoclonal antibody , 1996, Neuroscience Research.

[66]  Jeffrey C. Hall,et al.  Immunohistochemical localization of choline acetyltransferase during development and in Chats mutants of Drosophila melanogaster , 1987, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[67]  I. Salecker,et al.  Serotonin-immunoreactive neurons in the antennal lobes of the American cockroach Periplaneta americana: light- and electron-microscopic observations , 2004, Histochemistry.

[68]  D. Sattelle,et al.  Drosophila nervous system muscarinic acetylcholine receptor: transient functional expression and localization by immunocytochemistry. , 1993, Molecular pharmacology.

[69]  E. Gundelfinger,et al.  Expression of the ligand-binding nicotinic acetylcholine receptor subunit D alpha 2 in the Drosophila central nervous system. , 1994, Journal of neurobiology.

[70]  Reinhard F. Stocker,et al.  The organization of the chemosensory system in Drosophila melanogaster: a rewiew , 2004, Cell and Tissue Research.

[71]  R. Stocker,et al.  Larval chemosensory projections and invasion of adult afferents in the antennal lobe of Drosophila. , 1997, Journal of neurobiology.

[72]  Postembryonic development of γ‐aminobutyric acid‐like Immunoreactivity in the brain of the sphinx moth Manduca sexta , 1994 .

[73]  R. Stocker,et al.  Smell and Taste Perception in Drosophila melanogasterLarva: Toxin Expression Studies in Chemosensory Neurons , 1999, The Journal of Neuroscience.

[74]  D. Sattelle,et al.  Immunocytochemical mapping of a C-terminus anti-peptide antibody to the GABA receptor subunit, RDL in the nervous system of Drosophila melanogaster , 1996, Cell and Tissue Research.

[75]  Shubha V. Nayak,et al.  Primary sensory projections from the labella to the brain of Drosophila melanogaster Meigen (Diptera : Drosophilidae) , 1985 .