Quantifying behavior is crucial for many applications in neuroscience. Videography provides easy methods for the observation and recording of animal behavior in diverse settings, yet extracting particular aspects of a behavior for further analysis can be highly time consuming. In motor control studies, humans or other animals are often marked with reflective markers to assist with computer-based tracking, but markers are intrusive, and the number and location of the markers must be determined a priori. Here we present an efficient method for markerless pose estimation based on transfer learning with deep neural networks that achieves excellent results with minimal training data. We demonstrate the versatility of this framework by tracking various body parts in multiple species across a broad collection of behaviors. Remarkably, even when only a small number of frames are labeled (~200), the algorithm achieves excellent tracking performance on test frames that is comparable to human accuracy.

All-optical electrophysiology—spatially resolved simultaneous optical perturbation and measurement of membrane voltage—would open new vistas in neuroscience research. We evolved two archaerhodopsin-based voltage indicators, QuasAr1 and 2, which show improved brightness and voltage sensitivity, microsecond response times, and produce no photocurrent. We engineered a novel channelrhodopsin actuator, CheRiff, which shows improved light sensitivity and kinetics, and spectral orthogonality to the QuasArs. A co-expression vector, Optopatch, enabled crosstalk-free genetically targeted all-optical electrophysiology. In cultured neurons, we combined Optopatch with patterned optical excitation to probe back-propagating action potentials in dendritic spines, synaptic transmission, sub-cellular microsecond-timescale details of action potential propagation, and simultaneous firing of many neurons in a network. Optopatch measurements revealed homeostatic tuning of intrinsic excitability in human stem cell-derived neurons. In brain slice, Optopatch induced and reported action potentials and subthreshold events, with high signal-to-noise ratios. The Optopatch platform enables high-throughput, spatially resolved electrophysiology without use of conventional electrodes.

We have used endocytotic uptake of the styryl dye FM1-43 at synaptic terminals (Betz and Bewick, 1992) to study properties of individual synapses formed by axons of single hippocampal neurons in tissue culture. The distribution of values for probability of evoked transmitter release p estimated by dye uptake is continuous, with a preponderance of low p synapses and a broad spread of probabilities. We have validated this method by demonstrating that the optically estimated distribution of p at autapses in single-neuron microislands predicts, with no free parameters, the rate of blocking of NMDA responses by the noncompetitive antagonist MK-801 at the same synapses. Different synapses made by a single axon exhibited varying amounts of paired-pulse modulation; synapses with low p tended to be facilitated more than those with high p. The increment in release probability produced by increasing external calcium ion concentration also depended on a synapse's initial p value. The size of the recycling pool of vesicles was strongly correlated with p as well, suggesting that synapses with higher release probabilities had more vesicles. Finally, p values of neighboring synapses were correlated, indicating local interactions in the dendrite or axon, or both.

When hippocampal synapses in culture are pharmacologically silenced for several days, synaptic strength increases. The structural correlate of this change in strength is an increase in the size of the synapses, with all synaptic components--active zone, postsynaptic density, and bouton--becoming larger. Further, the number of docked vesicles and the total number of vesicles per synapse increases, although the number of docked vesicles per area of active zone is unchanged. In parallel with these anatomical changes, the physiologically measured size of the readily releasable pool (RRP) and the release probability are increased. Ultrastructural analysis of individual synapses in which the RRP was previously measured reveals that, within measurement error, the same number of vesicles are docked as are estimated to be in the RRP.

Synchronous 25-to 35-Hz were observed in local field potentials and unit activ in sensorimotor cortex of awake rhesus monkeys. The osatory epiode occurred often when the monkeys retrieved raisins from a KIliver board or from unseen locations uin somatosenor feedback; they occurred less often during perfnce of repetitive wrist flexion and extension movements. The amplitude, duration, and frequency of oscillations were not directly related to movement prameters In behaviors studied so far. The occurrence of the ollations was not nenty related to bursts of activity in foren musces, but cycle-triggered averages of eectromyograms revealed synchronous modulationin flexor and extensor muscles. The phase of the osilations changed continuously om te surface to the deeper layers ofthe cortex, reversing their polarity completely at depths exceeding 800 gAm. The oscillations could become synchronized over a distane of 14 mm mediolaterally in precentral cortex. Coherent osciations could also occur at pre-and postcentral sites separated by an estimated tangential intracortical dstance of20 mm.Activity ofsingle units was commonly seen to burst in synchrony with field potential llatins. These (3,5) and awake monkeys (6). Such oscillations occurred coherently at widely separated visual sites within and across the two hemispheres (3,5,7,8). Electroencephalogram (EEG) oscillations 'in the range of 20-50 Hz also occur in the motor and somatosensory cortex, especially during focused attention prior to motor acts (2, 9, 10). Pericruciate oscillations near 40 Hz occurred in cats (2), and in baboons and squirrel monkeys slower oscillations around 18 Hz were recorded at specific sites in the sensory hand areas and in area 5 (9). Scalp EEGs recorded from humans reportedly show an increase in 40-Hz signal during focused attention (10). Although the "40-Hz" oscillations are prevalent in the cortex, their function and the underlying mechanisms remain objects of speculation (11-13).The occurrence of coherent oscillations in single or multiunit activity in response to somatosensory stimulation or specific motor acts has not been reported to date, although one study reported that somatosensory stimulation disrupted rhythmic activity in single units (14). Here, we report the occurrence and properties of synchronous 25-to 35-Hz oscillations in the sensorimotor cortex of awake behaving monkeys. Oscillations in LFPs were most prominent during the performance of motor tasks that required attention and sensorimotor integration. The activities of some units separated by distances of up to 14 mm were found to be synchronized with each other and with the LFP oscillations.