Progress in experimental stroke and translational medicine could be accelerated by high-resolution in vivo imaging of disease progression in the mouse cortex. Here, we introduce optical microscopic methods that monitor brain injury progression using intrinsic optical scattering properties of cortical tissue. A multi-parametric Optical Coherence Tomography (OCT) platform for longitudinal imaging of ischemic stroke in mice, through thinned-skull, reinforced cranial window surgical preparations, is described. In the acute stages, the spatiotemporal interplay between hemodynamics and cell viability, a key determinant of pathogenesis, was imaged. In acute stroke, microscopic biomarkers for eventual infarction, including capillary non-perfusion, cerebral blood flow deficiency, altered cellular scattering, and impaired autoregulation of cerebral blood flow, were quantified and correlated with histology. Additionally, longitudinal microscopy revealed remodeling and flow recovery after one week of chronic stroke. Intrinsic scattering properties serve as reporters of acute cellular and vascular injury and recovery in experimental stroke. Multi-parametric OCT represents a robust in vivo imaging platform to comprehensively investigate these properties.

Objective Migraine is one of the most common and debilitating neurological conditions. Familial hemiplegic migraine type 1 (FHM1), a monogenic migraine subtype, is caused by gain-of-function of voltage-gated CaV2.1 calcium channels. FHM1 mice carry human pathogenic mutations in the α1A subunit of CaV2.1 channels and are highly susceptible to cortical spreading depression (CSD), the electrophysiologic event underlying migraine aura. To date, however, the mechanism underlying increased CSD/migraine susceptibility remains unclear. Methods We employed in vivo multiphoton microscopy of the genetically encoded Ca2+-indicator yellow cameleon to investigate synaptic morphology and [Ca2+]i in FHM1 mice. In order to study CSD-induced cerebral oligemia, we used in vivo laser speckle flowmetry and multimodal imaging. With electrophysiologic recordings we investigated the effect of the CaV2.1 gating modifier tert-butyl dihydroquinone on CSD in vivo. Results FHM1 mutations elevate neuronal [Ca2+]i and alter synaptic morphology as a mechanism for enhanced CSD susceptibility that we were able to normalize with a CaV2.1 gating modifier, in hyperexcitable FHM1 mice. At the synaptic level, axonal boutons were larger, and dendritic spines were predominantly mushroom type, which both provide a structural correlate for enhanced neuronal excitability. Resting neuronal [Ca2+]i was elevated in FHM1, with loss of compartmentalization between synapses and neuronal shafts. The percentage of calcium-overloaded neurons was increased. Neuronal [Ca2+]i surge during CSD was faster and larger, and post-CSD oligemia and hemoglobin desaturation were more severe in FHM1 brains. Interpretation Our findings provide a mechanism for enhanced CSD susceptibility in hemiplegic migraine. Abnormal synaptic Ca2+ homeostasis and morphology may contribute to chronic neurodegenerative changes as well as enhanced vulnerability to ischemia in migraineurs.

This paper describes a novel optical method for label-free quantitative imaging of cerebral blood flow (CBF) and intracellular motility (IM) in the rodent cerebral cortex. This method is based on a technique that integrates dynamic light scattering (DLS) and optical coherence tomography (OCT), named DLS-OCT. The technique measures both the axial and transverse velocities of CBF, whereas conventional Doppler OCT measures only the axial one. In addition, the technique produces a three-dimensional map of the diffusion coefficient quantifying nontranslational motions. In the DLS-OCT diffusion map, we observed high-diffusion spots, whose locations highly correspond to neuronal cell bodies and whose diffusion coefficient agreed with that of the motion of intracellular organelles reported in vitro in the literature. Therefore, the present method has enabled, for the first time to our knowledge, label-free imaging of the diffusion-like motion of intracellular organelles in vivo. As an example application, we used the method to monitor CBF and IM during a brief ischemic stroke, where we observed an induced persistent reduction in IM despite the recovery of CBF after stroke. This result supports that the IM measured in this study represent the cellular energy metabolism-related active motion of intracellular organelles rather than free diffusion of intracellular macromolecules.