The time required for the arrival of an ambulance crew and administration of first aid is critical to clinical outcome, particularly in the case of head injury victims requiring neuroprotective drugs following a car accident, falls, and assaults. Short response times of the medical team, together with proper treatment, can limit injury severity and even save a life before transportation to the nearest medical center. We present a comparative evaluation of five different neuroprotective drugs frequently used in intensive care and operating units in the early phase following traumatic brain injury (TBI): hypertonic saline (HTS), mannitol, morphine, melatonin, and minocycline. The effectiveness of these drugs in terms of changes in brain tissue morphology (cell organelle size, density, distribution, etc.) and biochemical tissue properties (chromophores’ content) was experimentally evaluated through analysis of the spectral reduced scattering and optical absorption coefficient parameters in the near-infrared (NIR) optical range (650 to 1000 nm). Experiments were conducted on anesthetized male mice subjected to a noninvasive closed head weight-drop model of focal TBI (n=50 and n=10 control) and monitored using an NIR diffuse reflectance spectroscopy system utilizing independent source–detector separation and location. After 10 min of baseline measurement, focal TBI was induced and measurements were conducted for 20 min. Subsequently, a neuroprotective drug was administrated and measurements were recorded for another 30 min. This work’s major findings are threefold: first, minocycline was found to improve hemodynamic outcome at the earliest time postinjury. Second, HTS decreased brain water content and inhibited the increase in intracranial pressure. Third, the efficacy of neuroprotective drugs can be monitored noninvasively with diffuse reflectance spectroscopy. The demonstrated ability to noninvasively detect cerebral physiological properties following early administration of neuroprotective drugs underlines the need for more extensive investigation of the combined use of clinical drugs in larger-scale preclinical experiments to find the most beneficial drug treatment for brain injury patients.

Recent advances in multichannel functional near-infrared spectroscopy (fNIRS) allow wide coverage of cortical areas while entailing the necessity to control family-wise errors (FWEs) due to increased multiplicity. Conventionally, the Bonferroni method has been used to control FWE. While Type I errors (false positives) can be strictly controlled, the application of a large number of channel settings may inflate the chance of Type II errors (false negatives). The Bonferroni-based methods are especially stringent in controlling Type I errors of the most activated channel with the smallest p value. To maintain a balance between Types I and II errors, effective multiplicity (M_eff) derived from the eigenvalues of correlation matrices is a method that has been introduced in genetic studies. Thus, we explored its feasibility in multichannel fNIRS studies. Applying the M_eff method to three kinds of experimental data with different activation profiles, we performed resampling simulations and found that M_eff was controlled at 10 to 15 in a 44-channel setting. Consequently, the number of significantly activated channels remained almost constant regardless of the number of measured channels. We demonstrated that the M_eff approach can be an effective alternative to Bonferroni-based methods for multichannel fNIRS studies.

It has been reported that a functional near-infrared spectroscopy (fNIRS) signal can be contaminated by extracerebral contributions. Many algorithms using multidistance separations to address this issue have been proposed, but their spatial separation performance has rarely been validated with simultaneous measurements of fNIRS and functional magnetic resonance imaging (fMRI). We previously proposed a method for discriminating between deep and shallow contributions in fNIRS signals, referred to as the multidistance independent component analysis (MD-ICA) method. In this study, to validate the MD-ICA method from the spatial aspect, multidistance fNIRS, fMRI, and laser-Doppler-flowmetry signals were simultaneously obtained for 12 healthy adult males during three tasks. The fNIRS signal was separated into deep and shallow signals by using the MD-ICA method, and the correlation between the waveforms of the separated fNIRS signals and the gray matter blood oxygenation level–dependent signals was analyzed. A three-way analysis of variance (signal depth×Hb kind×task) indicated that the main effect of fNIRS signal depth on the correlation is significant [F(1,1286)=5.34, p<0.05]. This result indicates that the MD-ICA method successfully separates fNIRS signals into spatially deep and shallow signals, and the accuracy and reliability of the fNIRS signal will be improved with the method.

The cytoarchitecture of the human brain is of great interest in diverse fields: neuroanatomy, neurology, neuroscience, and neuropathology. Traditional histology is a method that has been historically used to assess cell and fiber content in the ex vivo human brain. However, this technique suffers from significant distortions. We used a previously demonstrated optical coherence microscopy technique to image individual neurons in several square millimeters of en-face tissue blocks from layer II of the human entorhinal cortex, over 50  μm in depth. The same slices were then sectioned and stained for Nissl substance. We registered the optical coherence tomography (OCT) images with the corresponding Nissl stained slices using a nonlinear transformation. The neurons were then segmented in both images and we quantified the overlap. We show that OCT images contain information about neurons that is comparable to what can be obtained from Nissl staining, and thus can be used to assess the cytoarchitecture of the ex vivo human brain with minimal distortion. With the future integration of a vibratome into the OCT imaging rig, this technique can be scaled up to obtain undistorted volumetric data of centimeter cube tissue blocks in the near term, and entire human hemispheres in the future.

The optical monitoring of multiple single neuron activities requires high-throughput parallel acquisition of signals at millisecond temporal resolution. To this aim, holographic two-photon microscopy (2PM) based on spatial light modulators (SLMs) has been developed in combination with standard laser scanning microscopes. This requires complex coordinate transformations for the generation of holographic patterns illuminating the points of interest. We present a simpler and fully digital setup (SLM-2PM) which collects three-dimensional two-photon images by only exploiting the SLM. This configuration leads to an accurate placement of laser beamlets over small focal volumes, eliminating mechanically moving parts and making the system stable over long acquisition times. Fluorescence signals are diffraction limited and are acquired through a pixelated detector, setting the actual limit to the acquisition rate. High-resolution structural images were acquired by raster-scanning the sample with a regular grid of excitation focal volumes. These images allowed the selection of the structures to be further investigated through an interactive operator-guided selection process. Functional signals were collected by illuminating all the preselected points with a single hologram. This process is exemplified for high-speed (up to 1 kHz) two-photon calcium imaging on acute cerebellar slices.

Axonal injury and stress have long been thought to play a pathogenic role in a variety of neurodegenerative diseases. However, a model for studying single-cell axonal injury in mammalian cells and the processes of repair has not been established. The purpose of this study was to examine the response of neuronal growth cones to laser-induced axonal damage in cultures of embryonic rat hippocampal neurons and induced pluripotent stem cell (iPSC) derived human neurons. A 532-nm pulsed Nd:YVO₄ picosecond laser was focused to a diffraction limited spot at a precise location on an axon using a laser energy/power that did not rupture the cell membrane (subaxotomy). Subsequent time series images were taken to follow axonal recovery and growth cone dynamics. After laser subaxotomy, axons thinned at the damage site and initiated a dynamic cytoskeletal remodeling process to restore axonal thickness. The growth cone was observed to play a role in the repair process in both hippocampal and iPSC-derived neurons. Immunofluorescence staining confirmed structural tubulin damage and revealed initial phases of actin-based cytoskeletal remodeling at the damage site. The results of this study indicate that there is a repeatable and cross-species repair response of axons and growth cones after laser-induced damage.

Infrared neural stimulation (INS) is a neurostimulation modality that uses pulsed infrared light to evoke artifact-free, spatially precise neural activity with a noncontact interface; however, the technique has not been demonstrated in humans. The objective of this study is to demonstrate the safety and efficacy of INS in humans in vivo. The feasibility of INS in humans was assessed in patients (n=7) undergoing selective dorsal root rhizotomy, where hyperactive dorsal roots, identified for transection, were stimulated in vivo with INS on two to three sites per nerve with electromyogram recordings acquired throughout the stimulation. The stimulated dorsal root was removed and histology was performed to determine thermal damage thresholds of INS. Threshold activation of human dorsal rootlets occurred in 63% of nerves for radiant exposures between 0.53 and 1.23  J/cm². In all cases, only one or two monitored muscle groups were activated from INS stimulation of a hyperactive spinal root identified by electrical stimulation. Thermal damage was first noted at 1.09  J/cm² and a 2∶1 safety ratio was identified. These findings demonstrate the success of INS as a fresh approach for activating human nerves in vivo and providing the necessary safety data needed to pursue clinically driven therapeutic and diagnostic applications of INS in humans.

Computing microvascular cerebral blood flow (μCBF) in real cortical angiograms is challenging. Here, we investigated whether the use of Doppler optical coherence tomography (DOCT) flow measurements in individual vessel segments can help in reconstructing μCBF across the entire vasculature of a truncated cortical angiogram. A μCBF computational framework integrating DOCT measurements is presented. Simulations performed on a synthetic angiogram showed that the addition of DOCT measurements, especially close to large inflowing or outflowing vessels, reduces the impact of pressure boundary conditions and estimated vessel resistances resulting in a more accurate reconstruction of μCBF. Our technique was then applied to reconstruct microvascular flow distributions in the mouse cortex down to 660  μm by combining two-photon laser scanning microscopy angiography with DOCT.