Dissociated neurons of the pupal honeybee brain in cell culture

Kreissl, Sabine; Bicker, Gerd

doi:10.1007/bf01187116

Cited by 65 publications

(47 citation statements)

References 28 publications

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“…Cultured projection neurons or Kenyon cells do not retain their in vivo morphology. This is probably not a consequence of the labelling procedure, because it has been described previously for cultures of unlabelled Kenyon cells (Kreissl and Bicker, 1992) and unidentified antennal lobe neurons (Devaud et al, 1994;Kirchhof and Mercer, 1997). Also, in the present study branching patterns of labelled and unlabelled neurons within a given culture dish were indistinguishable.…”

Section: Identification and Culturing Of Projection Neuronssupporting

confidence: 76%

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Differential expression of voltage-sensitive K+ and Ca2+ currents in neurons of the honeybee olfactory pathway

Grünewald

2003

Journal of Experimental Biology

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The formation of olfactory memory is a multistage process that involves different areas of the insect brain. Olfactory conditioning of the proboscis extension reflex of the honeybee has proved to be a very powerful system for studying learningrelated neural mechanisms, enabling major neural elements of the olfactory and reward pathways in the honeybee brain to be identified (Erber et al., 1980;Hammer, 1993;Mauelshagen, 1993;Grünewald, 1999b;Hammer and Menzel, 1998) (for reviews, see Hammer, 1997;Menzel, 1999Menzel, , 2001) and several molecular pathways involved in this behaviour to be elucidated (e.g. Hildebrandt and Müller, 1995;Müller, 1996Müller, , 2000Grünbaum and Müller, 1998;Fiala et al., 1999) (for a review, see Menzel and Müller, 1996). Odors are perceived by sensillae on the honeybee antennae. The axons of the olfactory receptor neurons terminate within the primary olfactory neuropils of the honeybee brain, the antennal lobes. Olfactory information is processed here in a complex spatio-temporal fashion (Joerges et al., 1997;Stopfer et al., 1997;Sachse et al., 1999) and projection neurons transmit olfactory information from the antennal lobes to the lateral protocerebral lobes and the mushroom bodies within the protocerebrum (Homberg, 1984;Fonta et al., 1993;Abel et al., 2001). In the mushroom body calyces the projection neurons synapse onto mushroom body-intrinsic Kenyon cells, named after their discoverer (Kenyon, 1896). Olfactory information converges here with information from other sensory modalities, such as visual input in the mushroom body, and with reward-processing modulatory neurons from the suboesophageal ganglion (Erber et al., 1987;Hammer and Menzel, 1995). Intracellular recordings showed that odor applications induce complex spiking patterns in projection neurons, thus encoding olfactory and gustatory sensory information (Stopfer et al., 1997;Abel et al., 2001;Müller et al., 2002). The underlying ion channels of projection neurons have not yet been analysed, however, because the neurons could not be identified and distinguished from local interneurons, as is possible in other insects (Hayashi and Hildebrand, 1990;Oland and Hayashi, 1993

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Section: Identification and Culturing Of Projection Neuronssupporting

confidence: 76%

“…Kenyon cells and projection neurons were dissected and cultured following a modified protocol of Kreissl and Bicker (1992). After the staining procedure brains were removed from the head capsule and transferred into a preparation medium (Leibovitz L15, Gibco BRL; see below).…”

Section: Preparation Of Cell Cultures and Cell Identification In Vitromentioning

confidence: 99%

Differential expression of voltage-sensitive K+ and Ca2+ currents in neurons of the honeybee olfactory pathway

Grünewald

2003

Journal of Experimental Biology

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“…Previous studies of cultured Drosophila neurons of various stages from wild type and mutants allowed morphological and electrophysiological assessment, but without specific identification of the neurons beyond their regional origin (Salvaterra et al, 1987;Byerly and Leung, 1988;Solc and Aldrich, 1988;Wu, 1988;Li and Meinertzhagen, 1995;O'Dowd, 1995;Z hao and Wu, 1997). Primary cultures enriched for M B neurons have been established from brains of other insects (Kreissl and Bicker, 1992;Cayre et al, 1998), but the lack of tools for genetic manipulation limits the versatility of these systems. Acutely dissociated Drosophila M B neurons have been identified using M B-restricted lacZ reporter gene expression in combination with hypotonic loading of a fluorogenic substrate of ␤gal (Wright and Zhong, 1995;Delgado et al, 1998).…”

Section: The Kenyon Cell Culture Systemmentioning

confidence: 99%

The Steroid Hormone 20-Hydroxyecdysone Enhances Neurite Growth ofDrosophilaMushroom Body Neurons Isolated during Metamorphosis

1998

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Mushroom bodies (MBs) are symmetrically paired neuropils in the insect brain that are of critical importance for associative olfactory learning and memory. In Drosophila melanogaster, the MB intrinsic neurons (Kenyon cells) undergo extensive reorganization at the onset of metamorphosis. A phase of rapid axonal degeneration without cell death is followed by axonal regeneration. This re-elaboration occurs as levels of the steroid hormone 20-hydroxyecdysone (20E) are rising during the pupal stage. Based on the known role of 20E in directing many features of CNS remodeling during insect metamorphosis, we hypothesized that the outgrowth of MB axonal processes is promoted by 20E. Using a GAL4 enhancer trap line (201Y) that drives MB-restricted reporter gene expression, we identified Kenyon cells in primary cultures dissociated from early pupal CNS. Paired cultures derived from single brains isolated before the 20E pupal peak were incubated in medium with or without 20E for 2-4 d. Morphometric analysis demonstrated that MB neurons exposed to 20E had significantly greater total neurite length and branch number compared with that of MB neurons grown without hormone. The relationship between branch number and total neurite length remained constant regardless of hormone treatment in vitro, suggesting that 20E enhances the rate of outgrowth from pupal MB neurons in a proportionate manner and does not selectively increase neuritic branching. These results implicate 20E in enhancing axonal outgrowth of Kenyon cells to support MB remodeling during metamorphosis.

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“…Protocols for the cultivation of honey bee neuronal cells were developed about twenty years ago (Gascuel et al, 1991;Kreissl and Bicker, 1992;Devaud et al, 1994;Gascuel et al, 1994;). The original purpose at that time was to complement in vivo studies on the insect olfactory system with data from in vitro cell culture experiments.…”

Section: Isolation and Cultivation Of Primary Neuronal Cellsmentioning

confidence: 99%

Standard methods for cell cultures in Apis mellifera research

Genersch

Gisder

Hedtke

et al. 2013

Journal of Apicultural Research

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SummaryCell culture techniques are indispensable in most, if not all life science disciplines to date. Wherever appropriate cell culture models are lacking, scientific development is hampered. Unfortunately this has been and still is the case in honey bee research, because permanent honey bee cell lines have not so far been established. To overcome this hurdle, protocols for the cultivation of primary honey bee cells and of non-permanent honey bee cell lines have been developed. In addition, heterologous cell culture models for honey bee pathogens based on non-Apis insect cell lines have recently been developed. To further advance this progress and to encourage bee scientists to enter the field of cell biology based research, here we present protocols for the cultivation of honey bee primary cells and non-permanent cell lines, as well as hints for the cultivation of permanent insect cell lines suitable for honey bee research. Métodos estándar para cultivos celulares en Apis mellifera ResumenLas técnicas de cultivo celular son indispensables en la mayoría, si no en todas las disciplinas de ciencias de la vida hasta la fecha. Siempre que se carezca de modelos de cultivo celular apropiados, el desarrollo científico se ve obstaculizado. Desafortunadamente, esto ha sido y todavía es el caso de la investigación en la abeja de la miel, ya que hasta ahora, no se han establecido líneas celulares permanentes de abeja de la miel. Para superar este obstáculo, se han desarrollado protocolos para el cultivo de células primarias de la abeja de la miel y líneas celulares no permanentes de abejas. Además, también se han desarrollado recientemente modelos heterólogos de cultivo celular para patógenos de las abejas melíferas basados en líneas celulares de insectos que no pertenecen al género Apis. Para avanzar en este progreso y alentar a los científicos apícolas a entrar en el campo de la investigación basada en la biología celular, presentamos aquí los protocolos para el cultivo de células primarias de abejas y líneas celulares no permanentes, así como consejos para el cultivo de líneas celulares de insectos permanentes adecuadas para la investigación en la abeja de la miel.

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Dissociated neurons of the pupal honeybee brain in cell culture

Cited by 65 publications

References 28 publications

Differential expression of voltage-sensitive K+ and Ca2+ currents in neurons of the honeybee olfactory pathway

Differential expression of voltage-sensitive K+ and Ca2+ currents in neurons of the honeybee olfactory pathway

The Steroid Hormone 20-Hydroxyecdysone Enhances Neurite Growth ofDrosophilaMushroom Body Neurons Isolated during Metamorphosis

Standard methods for cell cultures in Apis mellifera research

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