This paper reports a new application of microfluidic devices in optogenetics and the study of learning behavior of model organisam. Due to the fully known genomic information and neuronal connections, Caenorhabditis elegans have been chosen as objects to study the preference establishment and their learning ability. The optogenetics-microfluidics manipulation platform provides a novel measurement method to quantify C.elegans behaviors and largely improve the experiment efficiency[1][2].
In our research, two channelrhodopsins, Chrimson and CoChR, are expressed in the pair of olfactory receptor neurons AWA and AWB, as well as the Ca2+ indicator, GEM-GECO protein. Modified worms were endued the artificial phenotype of heading for the blue light ray but avoiding the red light. Synchronic worms are selected and injected into the microfluidics devices. After operant conditioning and reinforcement training using laser excitation at specific wavelengths, their behaviors of moving forward or backward could be observed when worms semi-fixed in PDMS chambers. Corresponded neuron activities are indicated by the fluorescence intensity of GEM-GECO. Their responds to the odor of diacetyl and nonanone are also recorded. Data is collected to analyze their behavior deviation and adaptability under different conditioned stimulus.
The reprogramming chemotaxis responses of AWA, AWB were reported in 1997 [1]. It was also the first discovery of the determinant of olfactory preference. Based on the new optogenetics methods[3] and microfluidics devices[4], we study the probability to deviate their natural preference. Compared with conventional methods basically relied on chemical stimulation and behavioral statics[5][6], the optical activation of neurons is more fast and accurate. An overview of the responding mechanism in our neuron cell is shown in picture 1, and the schematic view of the microfluidics devices is in Figure 2, 3 and 4. The picture showing the position of olfactory receptor neurons is 5, and the excitation spectra of proteins in this research is shown in picture 6.
REFERENCES:
[1] Emily R. Troemel, Bruce E. Kimmel and Cornelia I. Bargmann, "Reprogramming chemotaxis responses: sensory neurons define olfactory preferences in C.elegans," J. Cell. 1997, Vol. 91, 161–169.
[2] Steven J. Husson, Alexander Gottschalk and Andrew M. Leifer, "Optogenetic manipulation of neural activity in C. elegans: From synapse to circuits and behavior," J. Cell. 2013. Vol. 105, 235-250.
[3] Nathan C Klapoetke, Yasunobu Murata et al, "Independent optical excitation of distinct neural populations," J. Nature Methods. 2014. Vol.11, No.3.
[4] Adriana San-Miguel and Hang Lu, "Microfluidics as a tool for C.elegans’ research." Wormbook. 2013.
[5] Ye, Hua-Yue, Bo-Ping Ye, and Da-Yong Wang. "Learning and learning choice in the nematode Caenorhabditis elegans." Neuroscience bulletin 22.6 (2006): 355-360.
[6] Ardiel, Evan L., and Catharine H. Rankin. "An elegant mind: learning and memory in Caenorhabditis elegans." Learning & Memory 17.4 (2010): 191-201.
