M. Bret Schneider

An optical neural interface: in vivo control of rodent motor cortex with integrated fiberoptic and optogenetic technology

Neural interface technology has made enormous strides in recent years but stimulating electrodes remain incapable of reliably targeting specific cell types (e.g. excitatory or inhibitory neurons) within neural tissue. This obstacle has major scientific and clinical implications. For example, there is intense debate among physicians, neuroengineers and neuroscientists regarding the relevant cell types recruited during deep brain stimulation (DBS); moreover, many debilitating side effects of DBS likely result from lack of cell-type specificity. We describe here a novel optical neural interface technology that will allow neuroengineers to optically address specific cell types in vivo with millisecond temporal precision. Channelrhodopsin-2 (ChR2), an algal light-activated ion channel we developed for use in mammals, can give rise to safe, light-driven stimulation of CNS neurons on a timescale of milliseconds. Because ChR2 is genetically targetable, specific populations of neurons even sparsely embedded within intact circuitry can be stimulated with high temporal precision. Here we report the first in vivo behavioral demonstration of a functional optical neural interface (ONI) in intact animals, involving integrated fiberoptic and optogenetic technology. We developed a solid-state laser diode system that can be pulsed with millisecond precision, outputs 20 mW of power at 473 nm, and is coupled to a lightweight, flexible multimode optical fiber, approximately 200 microm in diameter. To capitalize on the unique advantages of this system, we specifically targeted ChR2 to excitatory cells in vivo with the CaMKIIalpha promoter. Under these conditions, the intensity of light exiting the fiber ( approximately 380 mW mm(-2)) was sufficient to drive excitatory neurons in vivo and control motor cortex function with behavioral output in intact rodents. No exogenous chemical cofactor was needed at any point, a crucial finding for in vivo work in large mammals. Achieving modulation of behavior with optical control of neuronal subtypes may give rise to fundamental network-level insights complementary to what electrode methodologies have taught us, and the emerging optogenetic toolkit may find application across a broad range of neuroscience, neuroengineering and clinical questions.

Targeting and Readout Strategies for Fast Optical Neural Control In Vitro and In Vivo

Major obstacles faced by neuroscientists in attempting to unravel the complexity of brain function include both the heterogeneity of brain tissue (with a multitude of cell types present in vivo) and the high speed of brain information processing (with behaviorally relevant millisecond-scale electrical activity patterns). To address different aspects of these technical constraints, genetically targetable neural modulation tools have been developed by a number of groups (Zemelman et al., 2002; Banghart et al., 2004; Karpova et al., 2005; Lima and Miesenbock, 2005; Thompson et al., 2005; Chambers et al., 2006; Tan et al., 2006; Gorostiza et al., 2007; Lerchner et al., 2007; Szobota et al., 2007). One approach recently brought to neurobiology, combining both high speed and genetic targeting, is based on a family of fast light-responsive microbial opsins including halorhodopsins (e.g., NpHR) and channelrhodopsins (e.g., ChR2) (for review, see Zhang et al., 2007b). These microbial opsins are single-component transmembrane conductance regulators encompassing light sensitivity and fast membrane potential control within a single open reading frame, which can be used to achieve fast bidirectional control of specific cell types even in freely moving animals (Adamantidis et al., 2007; Zhang et al., 2007a). Although the basic functioning of these tools has been reviewed previously (Zhang et al., 2007b), here we describe a collection of targeting and readout strategies designed for rapid and flexible application of the microbial opsin system, and provide pointers to the relevant literature. Combinations of these multiple levels of targeting and readout define an evolving toolbox that may open up new possibilities for basic neuroscience investigation.

Tracking stem cell differentiation in the setting of automated optogenetic stimulation.

Membrane depolarization has been shown to play an important role in the neural differentiation of stem cells and in the survival and function of mature neurons. Here, we introduce a microbial opsin into ESCs and develop optogenetic technology for stem cell engineering applications, with an automated system for noninvasive modulation of ESC differentiation employing fast optogenetic control of ion flux. Mouse ESCs were stably transduced with channelrhodopsin‐2 (ChR2)‐yellow fluorescent protein and purified by fluorescence activated cell sorting (FACS). Illumination of resulting ChR2‐ESCs with pulses of blue light triggered inward currents. These labeled ESCs retained the capability to differentiate into functional mature neurons, assessed by the presence of voltage‐gated sodium currents, action potentials, fast excitatory synaptic transmission, and expression of mature neuronal proteins and neuronal morphology. We designed and tested an apparatus for optically stimulating ChR2‐ESCs during chronic neuronal differentiation, with high‐speed optical switching on a custom robotic stage with environmental chamber for automated stimulation and imaging over days, with tracking for increased expression of neural and neuronal markers. These data point to potential uses of ChR2 technology for chronic and temporally precise noninvasive optical control of ESCs both in vitro and in vivo, ranging from noninvasive control of stem cell differentiation to causal assessment of the specific contribution of transplanted cells to tissue and network function. STEM CELLS 2011;29:78–88

Characterization of a Novel Revolving Radiation Collimator

Introduction The ZAP-X is a novel self-contained and first-of-its-kind self-shielded therapeutic radiation device dedicated to brain and head and neck radiosurgery. By utilizing a 2.7-MV linear accelerator and incorporating a design in which a beam stop and major mechanical elements serve a radiation shielding function, the Zap-X does not typically require a radiation bunker. The unique collimator design of the Zap-X is especially critical to the performance of the overall system. The collimator consists of a shielded tungsten wheel oriented with its rotational axis perpendicular to the beam’s central axis; the goal of this design is to minimize radiation leakage. Beam selection is accomplished by rotating the wheel within its tungsten-shielded housing. We investigated radiation leakage from the Zap-X collimator to determine its compliance with internationally accepted standards using direct radiation measurements. Materials and methods To measure collimator leakage in the plane of the patient, equidistant measurement stations were defined in a plane perpendicular to the central beam axis (cax) 1 m from this axis (1 m from the radiation focal spot). To measure leakage alongside and adjacent to the accelerator, equidistant measurement stations were located 1 m from the cax along a line parallel to the cax in the plane of the collimator wheel and along a line parallel to the cax 90 degrees offset from the first line of stations. Results Radiation leakage emanating from the collimating head of the linear accelerator in the patient plane ranged between 4.0 and 10.4 mR. Radiation along the linear accelerator (1000 R delivered in the primary beam) varied between 1.7 and 6.8 mR and constituted between 0.00017% to 0.00068% of the primary beam. The former radiation originated from X-ray target leakage, while the latter is produced directly by the linear accelerator and both contributed to the overall leakage radiation that would reach a patient. Discussion Due to the large diameter of the Zap-X tungsten collimator wheel and the massive Zap-X tungsten cylindrical collimator shield, the overall patient leakage is 0.00104% of the primary beam at a 1-m distance from the beam central axis in the patient plane. Leakage radiation in the patient plane is limited by the International Electrotechnical Commission (IEC) to 0.1% of the total primary radiation. Radiation leakage along the linear accelerator and the collimator housing was determined to be 0.00068% of primary radiation intensity. This leakage value is lower than the 0.1% leakage limit stipulated by IEC by more than a factor of 100. Conclusions Typically, an MV radiation therapy system minimizes exposure by utilizing a combination of device and structural shielding. However, the Zap-X has been uniquely designed to minimize the need for structural shielding. Our results indicate radiation leakage from the collimator meets internationally accepted standards as defined by the IEC.

Self-Shielding Analysis of the Zap-X System

The Zap-X is a self-contained and first-of-its-kind self-shielded therapeutic radiation device dedicated to brain as well as head and neck stereotactic radiosurgery (SRS). By utilizing an S-band linear accelerator (linac) with a 2.7 megavolt (MV) accelerating potential and incorporating radiation-shielded mechanical structures, the Zap-X does not typically require a radiation bunker, thereby saving SRS facilities considerable cost. At the same time, the self-shielded features of the Zap-X are designed for more consistency of radiation protection, reducing the risk to radiation workers and others potentially exposed from a poorly designed or constructed radiotherapy vault. The hypothesis of the present study is that a radiosurgical system can be self-shielded such that it produces radiation exposure levels deemed safe to the public while operating under a full clinical workload. This study summarizes the Zap-X system shielding and found that the overall system radiation leakage values are reduced by a factor of 50 compared to the occupational radiation limit stipulated by the Nuclear Regulatory Commission (NRC) or agreement states. The goal of self-shielding is achieved under all but the most exceptional conditions for which additional room shielding or a larger restricted area in the vicinity of the Zap-X system would be required.