We can resolve multiple discrete features within a focal region of m spatial dimensions by first isolating each on the basis of n >/= 1 unique optical characteristics and then measuring their relative spatial coordinates. The minimum acceptable separation between features depends on the point-spread function in the (m + n)d-dimensional space formed by the spatial coordinates and the optical parameters, whereas the absolute spatial resolution is determined by the accuracy to which the coordinates can be measured. Estimates of each suggest that near-field fluorescence excitation microscopy/spectroscopy with molecular sensitivity and spatial resolution is possible.

Commentary: Inspired by my earlier work (see below) in single molecule imaging and the isolation of multiple exciton recombination sites within a single probe volume, here I proposed the principle which would eventually lead to PALM. Indeed, all methods of localization microscopy, including PALM, fPALM, PALMIRA, STORM, dSTORM, PAINT, GSDIM, etc. are specific embodiments of the general principle of single molecule isolation and localization I introduced here.

Locomotion requires coordinated motor activity throughout an animal's body. In both vertebrates and invertebrates, chains of coupled central pattern generators (CPGs) are commonly evoked to explain local rhythmic behaviors. In C. elegans, we report that proprioception within the motor circuit is responsible for propagating and coordinating rhythmic undulatory waves from head to tail during forward movement. Proprioceptive coupling between adjacent body regions transduces rhythmic movement initiated near the head into bending waves driven along the body by a chain of reflexes. Using optogenetics and calcium imaging to manipulate and monitor motor circuit activity of moving C. elegans held in microfluidic devices, we found that the B-type cholinergic motor neurons transduce the proprioceptive signal. In C. elegans, a sensorimotor feedback loop operating within a specific type of motor neuron both drives and organizes body movement.

Many animals possess mechanosensory neurons that fire when a limb nears the limit of its physical range, but the function of these proprioceptive limit detectors remains poorly understood. Here, we investigate a class of proprioceptors on the Drosophila leg called hair plates. Using calcium imaging in behaving flies, we find that a hair plate on the fly coxa (CxHP8) detects the limits of anterior leg movement. Reconstructing CxHP8 axons in the connectome, we found that they are wired to excite posterior leg movement and inhibit anterior leg movement. Consistent with this connectivity, optogenetic activation of CxHP8 neurons elicited posterior postural reflexes, while silencing altered the swing-to-stance transition during walking. Finally, we use comprehensive reconstruction of peripheral morphology and downstream connectivity to predict the function of other hair plates distributed across the fly leg. Our results suggest that each hair plate is specialized to control specific sensorimotor reflexes that are matched to the joint limit it detects. They also illustrate the feasibility of predicting sensorimotor reflexes from a connectome with identified proprioceptive inputs and motor outputs.

This paper identifies the prospects of using aphid species as ideal genetic model systems for the study of evolutionary developmental biology and genetic control of polyphenisms. The advantages and disadvantages of using aphids as genetic model organisms are discussed.

The increasing IC manufacturing cost encourages a business model where design houses outsource IC fabrication to remote foundries. Despite cost savings, this model exposes design houses to IC piracy as remote foundries can manufacture in excess to sell on the black market. Recent efforts in digital hardware security aim to thwart piracy by using XOR-based chip locking, cryptography, and active metering. To counter direct attacks and lower the exposure of unlocked circuits to the foundry, we introduce a multiplexor-based locking strategy that preserves test response allowing IC testing by an untrusted party before activation. We demonstrate a simple yet effective attack against a locked circuit that does not preserve test response, and validate the effectiveness of our locking strategy on IWLS 2005 benchmarks.

Biosensors exploit the remarkable specificity of biomolecular recognition to provide analytical tools that can measure the presence of a single molecular species in a complex mixture. A new strategy is emerging in the development of biosensor technologies: molecular-engineering techniques are being used to adapt the properties of proteins to simple, generic detector instrumentation, rather than adapting instruments to the unique requirements of a natural molecule.

There is considerable interest in the regulation of sensorimotor gating, since deficits in this process could play a critical role in the symptoms of schizophrenia and other psychiatric disorders. Sensorimotor gating is often studied in humans and rodents using the prepulse inhibition of the acoustic startle response (PPI) model, in which an acoustic prepulse suppresses behavioral output to a startle-inducing stimulus. However, the molecular and neural mechanisms underlying PPI are poorly understood. Here, we show that a regulatory pathway involving protein phosphatase 2A (PP2A), glycogen synthase kinase 3 beta (GSK3beta), and their downstream target, the M-type potassium channel, regulates PPI. Mice (Mus musculus) carrying a hypomorphic allele of Ppp2r5delta, encoding a regulatory subunit of PP2A, show attenuated PPI. This PPP2R5delta reduction increases the phosphorylation of GSK3beta at serine 9, which inactivates GSK3beta, indicating that PPP2R5delta positively regulates GSK3beta activity in the brain. Consistently, genetic and pharmacological manipulations that reduce GSK3beta function attenuate PPI. The M-type potassium channel subunit, KCNQ2, is a putative GSK3beta substrate. Genetic reduction of Kcnq2 also reduces PPI, as does systemic inhibition of M-channels with linopirdine. Importantly, both the GSK3 inhibitor 3-(2,4-dichlorophenyl)-4-(1-methyl-1H-indol-3-yl)1H-pyrrole-2,5-dione (SB216763) and linopirdine reduce PPI when directly infused into the medial prefrontal cortex (mPFC). Whole-cell electrophysiological recordings of mPFC neurons show that SB216763 and linopirdine have similar effects on firing, and GSK3 inhibition occludes the effects of M-channel inhibition. These data support a previously uncharacterized mechanism by which PP2A/GSK3beta signaling regulates M-type potassium channel activity in the mPFC to modulate sensorimotor gating.

In this review we discuss the current advances relating to structure determination from protein microcrystals with special emphasis on the newly developed method called MicroED. This method uses a transmission electron cryo-microscope to collect electron diffraction data from extremely small 3-dimensional (3D) crystals. MicroED has been used to solve the 3D structure of the model protein lysozyme to 2.9A resolution. As the method further matures, MicroED promises to offer a unique and widely applicable approach to protein crystallography using nanocrystals.

Expansion microscopy (ExM) is a method to expand biological specimens ~fourfold in each dimension by embedding in a hyper-swellable gel material. The expansion is uniform across observable length scales, enabling imaging of structures previously too small to resolve. ExM is compatible with any microscope and does not require expensive materials or specialized software, offering effectively sub-diffraction-limited imaging capabilities to labs that are not equipped to use traditional super-resolution imaging methods. Expanded specimens are ~99% water, resulting in strongly reduced optical scattering and enabling imaging of sub-diffraction-limited structures throughout specimens up to several hundred microns in (pre-expansion) thickness.