Advanced Circuit and Cellular Imaging Methods in Nonhuman Primates.

Novel genetically encoded tools and advanced microscopy methods have revolutionized neural circuit analyses in insects and rodents over the last two decades. Whereas numerous technical hurdles originally barred these methodologies from success in nonhuman primates (NHPs), current research has started to overcome those barriers. In some cases, methodological advances developed with NHPs have even surpassed their precursors. One such advance includes new ultra-large imaging windows on NHP cortex, which are larger than the entire rodent brain and allow analysis unprecedented ultra-large-scale circuits. NHP imaging chambers now remain patent for periods longer than a mouse's lifespan, allowing for long-term all-optical interrogation of identified circuits and neurons over timeframes that are relevant to human cognitive development. Here we present some recent imaging advances brought forth by research teams using macaques and marmosets. These include technical developments in optogenetics; voltage-, calcium- and glutamate-sensitive dye imaging; two-photon and wide-field optical imaging; viral delivery; and genetic expression of indicators and light-activated proteins that result in the visualization of tens of thousands of identified cortical neurons in NHPs. We describe a subset of the many recent advances in circuit and cellular imaging tools in NHPs focusing here primarily on the research presented during the corresponding mini-symposium at the 2019 Society for Neuroscience annual meeting.

Long-term all-optical interrogation of cortical neurons in awake-behaving nonhuman primates.

Whereas optogenetic techniques have proven successful in their ability to manipulate neuronal populations-with high spatial and temporal fidelity-in species ranging from insects to rodents, significant obstacles remain in their application to nonhuman primates (NHPs). Robust optogenetics-activated behavior and long-term monitoring of target neurons have been challenging in NHPs. Here, we present a method for all-optical interrogation (AOI), integrating optical stimulation and simultaneous two-photon (2P) imaging of neuronal populations in the primary visual cortex (V1) of awake rhesus macaques. A red-shifted channel-rhodopsin transgene (ChR1/VChR1 [C1V1]) and genetically encoded calcium indicators (genetically encoded calmodulin protein [GCaMP]5 or GCaMP6s) were delivered by adeno-associated viruses (AAVs) and subsequently expressed in V1 neuronal populations for months. We achieved optogenetic stimulation using both single-photon (1P) activation of neuronal populations and 2P activation of single cells, while simultaneously recording 2P calcium imaging in awake NHPs. Optogenetic manipulations of V1 neuronal populations produced reliable artificial visual percepts. Together, our advances show the feasibility of precise and stable AOI of cortical neurons in awake NHPs, which may lead to broad applications in high-level cognition and preclinical testing studies.

Saccadic eye movement metrics reflect surgical residents' fatigue.

OBJECTIVE: Little is known about the effects of surgical residents' fatigue on patient safety. We monitored surgical residents' fatigue levels during their call day using (1) eye movement metrics, (2) objective measures of laparoscopic surgical performance, and (3) subjective reports based on standardized questionnaires. BACKGROUND: Prior attempts to investigate the effects of fatigue on surgical performance have suffered from methodological limitations, including inconsistent definitions and lack of objective measures of fatigue, and nonstandardized measures of surgical performance. Recent research has shown that fatigue can affect the characteristics of saccadic (fast ballistic) eye movements in nonsurgical scenarios. Here we asked whether fatigue induced by time-on-duty (~24 hours) might affect saccadic metrics in surgical residents. Because saccadic velocity is not under voluntary control, a fatigue index based on saccadic velocity has the potential to provide an accurate and unbiased measure of the resident's fatigue level. METHODS: We measured the eye movements of members of the general surgery resident team at St. Joseph's Hospital and Medical Center (Phoenix, AZ) (6 males and 6 females), using a head-mounted video eye tracker (similar configuration to a surgical headlight), during the performance of 3 tasks: 2 simulated laparoscopic surgery tasks (peg transfer and precision cutting) and a guided saccade task, before and after their call day. Residents rated their perceived fatigue level every 3 hours throughout their 24-hour shift, using a standardized scale. RESULTS: Time-on-duty decreased saccadic velocity and increased subjective fatigue but did not affect laparoscopic performance. These results support the hypothesis that saccadic indices reflect graded changes in fatigue. They also indicate that fatigue due to prolonged time-on-duty does not result necessarily in medical error, highlighting the complicated relationship among continuity of care, patient safety, and fatigued providers. CONCLUSIONS: Our data show, for the first time, that saccadic velocity is a reliable indicator of the subjective fatigue of health care professionals during prolonged time-on-duty. These findings have potential impacts for the development of neuroergonomic tools to detect fatigue among health professionals and in the specifications of future guidelines regarding residents' duty hours.