Molecular correlates of memory phases and development of molecular tools to modify brain physiology in the honeybee.
Final Report Abstract
In this project, we gained knowledge on the neurotransmitter networks of the MB output lobes with emphasis on the glutamatergic neurotransmission and AmGluCl. Through collaborations, we could also work in parallel on projects requiring our expertise with the RNAi technique and in vivo electroporation. We could induce an RNAi response against AmGluClα. It was demonstrated by real-time PCR but we did not manage to characterise the reduction at the protein level. The development of the CLARITY technique to perform this analysis could not be achieved within the project. The technique was improved and it allows now to efficiently clear and perform synapsin detection in 30% of well preserved brains. Dr Bartels characterised extracellular currents induced by acetylcholine, glutamate and GABA in the vertical lobe. We attempted to analyse under different conditions the physiological properties of AmGluCl by mean of RNAi. Unfortunately our attempts to evaluate if glutamate induced currents depends on AmGluCl did not succeed. We circumvent the problem of the CLARITY technique by applying immuno-histological analysis with the Zinc-Chloride technique. In this manner we described the expression of AmGluCl in the brain and in particular in PCT neurons as it seems absent from KC. The project ended before we could evaluate the RNAi effect with this technique. However the results suggest that AmGluCl in PCT neurons participate to the inhibitory currents induced by glutamate in the lobes. Our in situ hybridisation and real-time PCR experiments on the neurotransmitter vesicular transporter completed our physiological and anatomical data. The description of some cholinergic KCs suggests that they are responsible of the Ach-induced activity within the vertical lobes. We could not detect vglut in KC, thus PCT neurons probably receive glutamatergic input from other cells. We still have to precise if some populations of PCT neurons might express vglut. There was also no monoaminergic and GABAergic KC, thus other neurotransmitters are expressed by KC. Our analysis of neuropeptides showed that the mRNA of MVP, IDL and TA are abundant in KC populations. Thus although the functional analysis developed unfortunately, we were successful in relating physiological and anatomical evidence. We could further develop the in vivo electroporation tools as planed. These results were published and the analysis of our data on in vivo electroporation will be the subject of a publication. Finally, our expertise in RNAi served in a study on pest control that was also published and we analysed and published results on a study on the LWL-opsin and its inhibition by RNAi.
Publications
- Characterisation of the RNAi response against the long-wavelength receptor of the honeybee. Insect Biochemistry and Molecular Biology – 43:959-969.
Leboulle G., Niggebrügge C., Rößler R., Menzel R., Hempel de Ibarra N.
(See online at https://doi.org/10.1016/j.ibmb.2013.07.006) - Honeybee promoter sequences for targeted gene expression. Insect Molecular Biology – 22(4):399-410.
Schulte C., Leboulle G., Otte M., Grünewald B., Gehne N., Beye M.
(See online at https://doi.org/10.1111/imb.12031) - Large scale RNAi screen in Tribolium reveals novel target genes for pest control and the proteasome as prime target. BMC Genomics (2015) Sep 3;16:674.
Ulrich J., Dao V.A., Majumdar U., Schmitt-Engel C., Schwirz J., Schultheis D., Ströhlein N., Troelenberg N., Grossmann D., Richter T., Dönitz J., Gerischer L., Leboulle G., Vilcinskas A., Stanke M., Bucher G.
(See online at https://doi.org/10.1186/s12864-015-1880-y) - Identification of the neurotransmitter profile of AmFoxP expressing neurons in the honeybee brain using double-label in situ hybridization. BMC neuroscience, Vol. 19. 2018, Article number: 69.
Schatton, A., Agoro, J., Mardink, J., Leboulle, G., & Scharff, C.
(See online at https://doi.org/10.1186/s12868-018-0469-1)
