Advanced Imaging Core

National Institute On Deafness And Other Communication Disorders

Investigators

Linked publications & trials

Abstract

Ronald S. Petralia and Ya-Xian Wang have run the Advanced Imaging Core facility of NIDCD since its inception in October 2011. This core is dedicated primarily to collaborating with and training other scientists in NIDCD in studies utilizing transmission electron microscopy (TEM). And we have presented an annual workshop on TEM as part of the EARssentials program, prior to the pandemic. Some work also is done with other NIH institutes. In addition, Ronald Petralia regularly reviews papers for professional journals and reviews many various documents for NIDCD PIs, reviews grants, writes review papers and letters of recommendation, serves on several NIH committees, and helps coordinate lab+office safety for NIDCD (including annual inspections and reports), and just previous to this fiscal year, performed animal facility inspections as part of his duties in the ACUC. Ronald Petralia and Ya-Xian Wang also recently submitted an invited review of immunogold techniques to Frontiers in Neuroanatomy. We had two major upgrades to the JEOL JEM-2100 TEM in 2019, including a new Gatan OneView camera with greater sensitivity and drift correction, and a turbo pump. The turbo pump is more reliable and is less sensitive to humidity. With the addition of the turbo pump, we were able to look at our first cryo-EM specimen this past year and work continues to facilitate the cryo-EM workflow on this microscope. Additionally, the installation of our second electron microscope that will be dedicated largely to cryo-EM began this past November; it is a Thermo Fisher/FEI Titan Halo 300 kV TEM. Dennis Winkler, who joined NIDCD and this core in June 2019, is a top expert on cryo-EM; he is overseeing the installation of the Halo and will be responsible for cryo-EM for both TEMs. Although there have been some lingering issues with the room renovation and the building house chilled water systems, we still expect the microscope to come online in the beginning of 2022. Dennis Winkler, Thermo Fisher/FEI (the Halos manufacturer), NIH's facility management team (ORF, DFOM, and building engineers) and the renovation team are working to ensure that all is right with the renovation and scope installation. Also, Dennis Winkler has begun working with members of Dr. Friedman's and Dr. Kachar's labs to train personnel in cryo-EM. As his former NIAMS lab is still winding down, he still offers assistance with finishing manuscripts, reviewing papers, and, as the most knowledgeable on the lab space and its equipment, helps in the decommissioning of equipment and the transitioning of the space to others in NIAMS and at NIH. Dennis Winkler continues to be the de facto manager for the JEOL 2200 electron microscope in Building 50 as NIAMS has not appointed a new manager and NIDCD is using this scope for cryo-EM work and training. Julia Sokolova joined the core on half-time at the beginning of 2021; she works directly for the Friedman lab for the other half of her time. Julia has a strong background in TEM of microorganisms. Both the work for the core and for Dr. Friedman's lab involve similar multiple methods in TEM, and Ron Petralia and Ya-Xian Wang have been actively training Julia on TEM methods as used for the study of auditory structures. NIDCD: Tom Friedman/Inna Belyantseva lab: We are currently involved in a collaboration, although this has slowed due to the pandemic situation. Also, this core continues to advise Inna Belyantseva on TEM techniques, to optimize ultrastructural studies of hair cells and stereocilia. We also assist Inna with tissue preparation, e.g., freeze substitution and ultramicrotomy. Julia Sokolova joined the Friedman lab for half-time and now works with Inna on TEM projects as we continue to train her. Tom Friedman/Takushi Miyoshi lab: This core has started its inaugural cryo-EM collaboration with Drs. Friedman and Miyoshi. The project will examine methods to detect abnormal functions of deafness gene variants in hair cell stereocilia Isabelle Roux group (laboratory of former SD, Andrew Griffith): We began a major study on the structure and function of the endolymphatic sac, with preliminary work done in March 2020; this was halted abruptly due to the COVID-19 pandemic. This has continued through much of FY21. Katie Kindt lab: We recently began planning a new study on zebrafish hair cells that will involve Julia, Ron, and Ya-Xian. Ron and Ya-Xian will be training Julia in the study of TEM on these fish over the next few months. Bechara Kachar lab: We provide extensive training for Ranjan Sengupta and Felipe Montecinos on TEM use and TEM preparation methods for both traditional and cryo-EM. Additional assistance to the lab includes review of manuscripts. Other NIH institutes: NINDS: We have a study with the Lu lab on Neuroligin 2 that is in final preparation for submission (Han et al.; delayed by the COVID-19 pandemic). NICHD: We published a paper for the Hoffman lab, that describes a novel structure associated with aging dementia in DPP6-KO mice, at the beginning of FY21; Ron Petralia is co-first author (Lin et al., 2020). We also recently published a study on microtubule transport profile of Kv4.2 in axons versus dendrites (Bellotti et al., 2021). Another study examining the association of Cav2.3 and Kv4.2 channels is in revision for the journal, Cell Reports (Murphy et al.). Then, we have a paper in final preparation that is a follow up study on the novel structure associated with aging dementia in DPP6-KO mice that we published in 2020. In addition, we have 2 studies on zebrafish neuromast development, with the former lab of Katie Drerup; currently, the status of these is unknown. NIA: We have been working with Pamela Yao/Dimitrios Kapogiannis on studies related to neuronal mitochondria and exosomes. Ron Petralia was invited to write a review paper on invaginating processes in synapses for a special topics issue of Frontiers in Synaptic Neuroscience, and this was published recently in collaboration with Pamela Yao (Petralia et al., 2021). We currently are working on a project on mitochondrial extracellular vesicles, and it involves Ron and Ya-Xian, as well as Julia Sokolova. NIMH: We have recently published 2 papers with the lab of Zheng Li including one on BAD-BAX-caspase-3 cascade control of synaptic vesicle pools via autophagy in The Journal of Neuroscience (Gu et al., 2021, J. Neurosci. 41:1174-1190), and we published a study on mitochondrial fission and dysbindin-1 (Zhao et al., 2021; Molec. Psych., in press). Then we have a study on mitophagy in the amygdala and hippocampus associated with social anxiety, currently in press for the journal, Neuron (Duan et al., 2021). NHLBI: We had a paper published on a special myosin found in Purkinje spines with the lab of John Hammer (additional NIDCD staff, Melanie Barzik and Tom Friedman, are authors on this; Alexander et al., 2021, The FASEB Journal. 2021;35:e21092). NHGRI: We continue to work periodically on studies with Nahid Tayebi/Ellen Sidransky on Parkinson and Gaucher diseases. NIAMS: We continue working with Elif Eren and Norman Watts of the Protein Expression Lab, assisting them with their protein complex studies. There is the potential for collaboration in these studies using the above-mentioned Halo as their access to EM equipment may diminish in the future.