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The role of dopamine in Drosophila larval classical olfactory conditioning.

Selcho M, Pauls D, Han KA, Stocker RF, Thum AS - PLoS ONE (2009)

Bottom Line: Single cell analysis suggests that dopaminergic neurons do not directly connect gustatory input in the larval suboesophageal ganglion to olfactory information in the mushroom bodies.We found that dopamine receptors are highly enriched in the mushroom bodies and that aversive and appetitive olfactory learning is strongly impaired in dopamine receptor mutants.Genetically interfering with dopaminergic signaling supports this finding, although our data do not exclude on naïve odor and sugar preferences of the larvae.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Fribourg, Fribourg, Switzerland.

ABSTRACT
Learning and memory is not an attribute of higher animals. Even Drosophila larvae are able to form and recall an association of a given odor with an aversive or appetitive gustatory reinforcer. As the Drosophila larva has turned into a particularly simple model for studying odor processing, a detailed neuronal and functional map of the olfactory pathway is available up to the third order neurons in the mushroom bodies. At this point, a convergence of olfactory processing and gustatory reinforcement is suggested to underlie associative memory formation. The dopaminergic system was shown to be involved in mammalian and insect olfactory conditioning. To analyze the anatomy and function of the larval dopaminergic system, we first characterize dopaminergic neurons immunohistochemically up to the single cell level and subsequent test for the effects of distortions in the dopamine system upon aversive (odor-salt) as well as appetitive (odor-sugar) associative learning. Single cell analysis suggests that dopaminergic neurons do not directly connect gustatory input in the larval suboesophageal ganglion to olfactory information in the mushroom bodies. However, a number of dopaminergic neurons innervate different regions of the brain, including protocerebra, mushroom bodies and suboesophageal ganglion. We found that dopamine receptors are highly enriched in the mushroom bodies and that aversive and appetitive olfactory learning is strongly impaired in dopamine receptor mutants. Genetically interfering with dopaminergic signaling supports this finding, although our data do not exclude on naïve odor and sugar preferences of the larvae. Our data suggest that dopaminergic neurons provide input to different brain regions including protocerebra, suboesophageal ganglion and mushroom bodies by more than one route. We therefore propose that different types of dopaminergic neurons might be involved in different types of signaling necessary for aversive and appetitive olfactory memory formation respectively, or for the retrieval of these memory traces. Future studies of the dopaminergic system need to take into account such cellular dissociations in function in order to be meaningful.

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Single Cell Staining of Potentially Dopaminergic Neurons.The left column shows the projection pattern of each cell type. The other columns represent higher magnifications of neuropile regions (magenta; A″, B″, C′ and G″ show single confocal sections) innervated by the respective cell type (green). (A–A′″) Sagittal view of the innervation pattern of the DL2-1 cell type. DL2-1 projects to both calyces (ca; arrow A″ and A′″) and ramifies in the dorsal protocerebrum (dp). The posterior pedunculi (ped) show innervations (arrowhead A″). (B–B′″) Two neurons of the DM3 cell type one of which is stained much weaker (arrowhead). (B′) The axon crossing the midline terminates at the contralateral ca (arrow). The ipsilateral ca and dp are innervated (B″ and B′″). (C–C″) The ipsilateral projecting DL2-2 cell type innervates the basolateral/mediolateral ca (arrow C′). Ramifications in the dorsolateral protocerebrum (dlp) were observed (C′–C′″). DL2-2 bifurcates around the ped (C′ and C″) and reaches the neuropile lateral to the posterior part of the medial lobe (ml; arrow C″). (D–D′″) DL1-6 shows the characteristic dorsomedial protocerebrum (dmp) innervation. A small axon projects through the vertical lobe (vl) without any ramifications (arrowhead D′). The basomedial protocerebra (bmp) are innervated. (E–E″) DM4 innervates the dp around the vl. (E′) Two axons project through the lobe (arrowhead). A dot-like terminal is observed in the lateral appendix (arrow). (E″) The protocerebrum lateral to the ml is innervated. (F–F″) The innervation of DL2-3 is restricted to the basal protocerebrum (bp). The primary neurite bifurcates in the posterior bmp (arrow F′). One secondary neurite innervates the bp and posterior sog (F′). The other is sent to the contralateral side and ramifies in the bp (F″). (G–G″) DM5 innervates the anterior basolateral protocerebrum (blp) ventral to the vl. (G″) Single section of the posterior blp and posterior sog. Scale bars: left column 50 µm, rest 25 µm.
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pone-0005897-g006: Single Cell Staining of Potentially Dopaminergic Neurons.The left column shows the projection pattern of each cell type. The other columns represent higher magnifications of neuropile regions (magenta; A″, B″, C′ and G″ show single confocal sections) innervated by the respective cell type (green). (A–A′″) Sagittal view of the innervation pattern of the DL2-1 cell type. DL2-1 projects to both calyces (ca; arrow A″ and A′″) and ramifies in the dorsal protocerebrum (dp). The posterior pedunculi (ped) show innervations (arrowhead A″). (B–B′″) Two neurons of the DM3 cell type one of which is stained much weaker (arrowhead). (B′) The axon crossing the midline terminates at the contralateral ca (arrow). The ipsilateral ca and dp are innervated (B″ and B′″). (C–C″) The ipsilateral projecting DL2-2 cell type innervates the basolateral/mediolateral ca (arrow C′). Ramifications in the dorsolateral protocerebrum (dlp) were observed (C′–C′″). DL2-2 bifurcates around the ped (C′ and C″) and reaches the neuropile lateral to the posterior part of the medial lobe (ml; arrow C″). (D–D′″) DL1-6 shows the characteristic dorsomedial protocerebrum (dmp) innervation. A small axon projects through the vertical lobe (vl) without any ramifications (arrowhead D′). The basomedial protocerebra (bmp) are innervated. (E–E″) DM4 innervates the dp around the vl. (E′) Two axons project through the lobe (arrowhead). A dot-like terminal is observed in the lateral appendix (arrow). (E″) The protocerebrum lateral to the ml is innervated. (F–F″) The innervation of DL2-3 is restricted to the basal protocerebrum (bp). The primary neurite bifurcates in the posterior bmp (arrow F′). One secondary neurite innervates the bp and posterior sog (F′). The other is sent to the contralateral side and ramifies in the bp (F″). (G–G″) DM5 innervates the anterior basolateral protocerebrum (blp) ventral to the vl. (G″) Single section of the posterior blp and posterior sog. Scale bars: left column 50 µm, rest 25 µm.

Mentions: The maximally eight stained neurons of the DL1 cluster could be categorized into six different types (DL1-1–DL1-6). As the DL1-4 type was hit twice as often as the others (Table 2), we speculated that it was represented twice in the DL1 cluster. The DL2 cluster, which consisted of about six neurons, seemed to be organized in three different cell types. Interestingly, two types occurred more often in our flp-out clones; therefore we speculated that these were also represented twice per hemisphere (DL2-2 and DL2-3; Table 2). Yet, we cannot exclude that we missed one cell type in the DL2 cluster. For the DM cluster, we were able to identify five different types of neurons. However, due to the weak expression in other cells, we were not able to classify all of these neurons; here the flp-out technique displayed obvious limitations. In the SM clusters we identified all TH-GAL4 positive neurons, which belonged to four different types. Multi-cell clones suggested that one type occurred four times (SM1-2; Table 2). For the SL cluster we described two types. Again one type emerged more often in our analysis, suggesting that it comprises at least two copies (SL2; Table 2). In total, we identified a comprehensive set of TH-GAL4 positive neurons, leaving only weakly labelled neurons unidentified due to the limitations of the flp-out technique (Figures 5, 6 and 7). At least 19 different types of TH-GAL4 positive, mostly paired neurons were found to innervate the brain, for example the mbs; twelve types innervated the sog (Figures 5, 6 and 7). Remarkably, none of these neurons innervated both the mb and GRN input region of the sog. Next, we focused on candidates potentially involved in aversive and appetitive olfactory learning, i.e. neurons having their cell body in the hemispheres, sog or thoracic ganglion. The terminology used refers to their association with the DL, DM, S or T clusters, followed by a further subdivision into different types. The following sections first describes cells innervating the mb lobes and/or the pedunculi (Figure 5), then those sending their branches onto the calyces (Figure 6A–6C), and finally the neurons that may have a limited mb projection (Figure 6D–6G).


The role of dopamine in Drosophila larval classical olfactory conditioning.

Selcho M, Pauls D, Han KA, Stocker RF, Thum AS - PLoS ONE (2009)

Single Cell Staining of Potentially Dopaminergic Neurons.The left column shows the projection pattern of each cell type. The other columns represent higher magnifications of neuropile regions (magenta; A″, B″, C′ and G″ show single confocal sections) innervated by the respective cell type (green). (A–A′″) Sagittal view of the innervation pattern of the DL2-1 cell type. DL2-1 projects to both calyces (ca; arrow A″ and A′″) and ramifies in the dorsal protocerebrum (dp). The posterior pedunculi (ped) show innervations (arrowhead A″). (B–B′″) Two neurons of the DM3 cell type one of which is stained much weaker (arrowhead). (B′) The axon crossing the midline terminates at the contralateral ca (arrow). The ipsilateral ca and dp are innervated (B″ and B′″). (C–C″) The ipsilateral projecting DL2-2 cell type innervates the basolateral/mediolateral ca (arrow C′). Ramifications in the dorsolateral protocerebrum (dlp) were observed (C′–C′″). DL2-2 bifurcates around the ped (C′ and C″) and reaches the neuropile lateral to the posterior part of the medial lobe (ml; arrow C″). (D–D′″) DL1-6 shows the characteristic dorsomedial protocerebrum (dmp) innervation. A small axon projects through the vertical lobe (vl) without any ramifications (arrowhead D′). The basomedial protocerebra (bmp) are innervated. (E–E″) DM4 innervates the dp around the vl. (E′) Two axons project through the lobe (arrowhead). A dot-like terminal is observed in the lateral appendix (arrow). (E″) The protocerebrum lateral to the ml is innervated. (F–F″) The innervation of DL2-3 is restricted to the basal protocerebrum (bp). The primary neurite bifurcates in the posterior bmp (arrow F′). One secondary neurite innervates the bp and posterior sog (F′). The other is sent to the contralateral side and ramifies in the bp (F″). (G–G″) DM5 innervates the anterior basolateral protocerebrum (blp) ventral to the vl. (G″) Single section of the posterior blp and posterior sog. Scale bars: left column 50 µm, rest 25 µm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2690826&req=5

pone-0005897-g006: Single Cell Staining of Potentially Dopaminergic Neurons.The left column shows the projection pattern of each cell type. The other columns represent higher magnifications of neuropile regions (magenta; A″, B″, C′ and G″ show single confocal sections) innervated by the respective cell type (green). (A–A′″) Sagittal view of the innervation pattern of the DL2-1 cell type. DL2-1 projects to both calyces (ca; arrow A″ and A′″) and ramifies in the dorsal protocerebrum (dp). The posterior pedunculi (ped) show innervations (arrowhead A″). (B–B′″) Two neurons of the DM3 cell type one of which is stained much weaker (arrowhead). (B′) The axon crossing the midline terminates at the contralateral ca (arrow). The ipsilateral ca and dp are innervated (B″ and B′″). (C–C″) The ipsilateral projecting DL2-2 cell type innervates the basolateral/mediolateral ca (arrow C′). Ramifications in the dorsolateral protocerebrum (dlp) were observed (C′–C′″). DL2-2 bifurcates around the ped (C′ and C″) and reaches the neuropile lateral to the posterior part of the medial lobe (ml; arrow C″). (D–D′″) DL1-6 shows the characteristic dorsomedial protocerebrum (dmp) innervation. A small axon projects through the vertical lobe (vl) without any ramifications (arrowhead D′). The basomedial protocerebra (bmp) are innervated. (E–E″) DM4 innervates the dp around the vl. (E′) Two axons project through the lobe (arrowhead). A dot-like terminal is observed in the lateral appendix (arrow). (E″) The protocerebrum lateral to the ml is innervated. (F–F″) The innervation of DL2-3 is restricted to the basal protocerebrum (bp). The primary neurite bifurcates in the posterior bmp (arrow F′). One secondary neurite innervates the bp and posterior sog (F′). The other is sent to the contralateral side and ramifies in the bp (F″). (G–G″) DM5 innervates the anterior basolateral protocerebrum (blp) ventral to the vl. (G″) Single section of the posterior blp and posterior sog. Scale bars: left column 50 µm, rest 25 µm.
Mentions: The maximally eight stained neurons of the DL1 cluster could be categorized into six different types (DL1-1–DL1-6). As the DL1-4 type was hit twice as often as the others (Table 2), we speculated that it was represented twice in the DL1 cluster. The DL2 cluster, which consisted of about six neurons, seemed to be organized in three different cell types. Interestingly, two types occurred more often in our flp-out clones; therefore we speculated that these were also represented twice per hemisphere (DL2-2 and DL2-3; Table 2). Yet, we cannot exclude that we missed one cell type in the DL2 cluster. For the DM cluster, we were able to identify five different types of neurons. However, due to the weak expression in other cells, we were not able to classify all of these neurons; here the flp-out technique displayed obvious limitations. In the SM clusters we identified all TH-GAL4 positive neurons, which belonged to four different types. Multi-cell clones suggested that one type occurred four times (SM1-2; Table 2). For the SL cluster we described two types. Again one type emerged more often in our analysis, suggesting that it comprises at least two copies (SL2; Table 2). In total, we identified a comprehensive set of TH-GAL4 positive neurons, leaving only weakly labelled neurons unidentified due to the limitations of the flp-out technique (Figures 5, 6 and 7). At least 19 different types of TH-GAL4 positive, mostly paired neurons were found to innervate the brain, for example the mbs; twelve types innervated the sog (Figures 5, 6 and 7). Remarkably, none of these neurons innervated both the mb and GRN input region of the sog. Next, we focused on candidates potentially involved in aversive and appetitive olfactory learning, i.e. neurons having their cell body in the hemispheres, sog or thoracic ganglion. The terminology used refers to their association with the DL, DM, S or T clusters, followed by a further subdivision into different types. The following sections first describes cells innervating the mb lobes and/or the pedunculi (Figure 5), then those sending their branches onto the calyces (Figure 6A–6C), and finally the neurons that may have a limited mb projection (Figure 6D–6G).

Bottom Line: Single cell analysis suggests that dopaminergic neurons do not directly connect gustatory input in the larval suboesophageal ganglion to olfactory information in the mushroom bodies.We found that dopamine receptors are highly enriched in the mushroom bodies and that aversive and appetitive olfactory learning is strongly impaired in dopamine receptor mutants.Genetically interfering with dopaminergic signaling supports this finding, although our data do not exclude on naïve odor and sugar preferences of the larvae.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Fribourg, Fribourg, Switzerland.

ABSTRACT
Learning and memory is not an attribute of higher animals. Even Drosophila larvae are able to form and recall an association of a given odor with an aversive or appetitive gustatory reinforcer. As the Drosophila larva has turned into a particularly simple model for studying odor processing, a detailed neuronal and functional map of the olfactory pathway is available up to the third order neurons in the mushroom bodies. At this point, a convergence of olfactory processing and gustatory reinforcement is suggested to underlie associative memory formation. The dopaminergic system was shown to be involved in mammalian and insect olfactory conditioning. To analyze the anatomy and function of the larval dopaminergic system, we first characterize dopaminergic neurons immunohistochemically up to the single cell level and subsequent test for the effects of distortions in the dopamine system upon aversive (odor-salt) as well as appetitive (odor-sugar) associative learning. Single cell analysis suggests that dopaminergic neurons do not directly connect gustatory input in the larval suboesophageal ganglion to olfactory information in the mushroom bodies. However, a number of dopaminergic neurons innervate different regions of the brain, including protocerebra, mushroom bodies and suboesophageal ganglion. We found that dopamine receptors are highly enriched in the mushroom bodies and that aversive and appetitive olfactory learning is strongly impaired in dopamine receptor mutants. Genetically interfering with dopaminergic signaling supports this finding, although our data do not exclude on naïve odor and sugar preferences of the larvae. Our data suggest that dopaminergic neurons provide input to different brain regions including protocerebra, suboesophageal ganglion and mushroom bodies by more than one route. We therefore propose that different types of dopaminergic neurons might be involved in different types of signaling necessary for aversive and appetitive olfactory memory formation respectively, or for the retrieval of these memory traces. Future studies of the dopaminergic system need to take into account such cellular dissociations in function in order to be meaningful.

Show MeSH
Related in: MedlinePlus