SignificanceThere are many neuroscience questions that can be answered by a high-resolution functional brain imaging system. Such a system would require the capability to visualize vasculature and measure neural activity by imaging the entire brain continually and in rapid succession in order to capture hemodynamic changes. Utilizing optical excitation and acoustic detection, photoacoustic technology enables label-free quantification of changes in endogenous chromophores, such as oxyhemoglobin, deoxyhemoglobin, and total hemoglobin.AimOur aim was to develop a sufficiently high-resolution, fast frame-rate, and wide field-of-view (FOV) photoacoustic microscopy (PAM) system for the purpose of imaging vasculature and hemodynamics in a rat brain.ApproachAlthough the most PA microscopy systems use raster scanning (or less commonly Lissajous scanning), we have developed a simple-to-implement laser scanning optical resolution PAM system with spiral scanning (which we have named “spiral laser scanning photoacoustic microscopy” or sLS-PAM) to acquire an 18 mm diameter image at fast frame rate (more than 1 fps). Such a system is designed to permit continuous rat brain imaging without the introduction of photobleaching artifacts.ConclusionWe demonstrated the functional imaging capability of the sLS-PAM system by imaging cerebral hemodynamics in response to whisker and electrical stimulation and used it for vascular imaging of a modeled brain injury. We believe that we have demonstrated the development of a simple-to-implement PAM system, which could become an affordable functional neuroimaging tool for researchers.
We have developed a fast, wide field-of-view, simple-to-implement laser scanning photoacoustic microscopy (PAM) system that is capable of imaging of the whole rat brain in < 1s. The system utilizes galvo scanners and a novel scanning scheme, that can counter the trade off between imaging speed and scanning area. We performed in-vivo imaging of rat brain to demonstrate the imaging capability of the system.
Current functional imaging techniques, such as functional magnetic resonance imaging, rely upon activity-induced blood flood changes to neurons. This indirect measurement of neuronal activity inherently limits image resolution and specificity, however, advances in transgenic technology and Photoacoustic (PA) methodology have offered new solutions. We previously demonstrated using PA imaging in conjunction with a Fos-LacZ rat model to map activated neurons through selective formation of PA-active X-gal product within Fos-expressing neurons. While we have previously used intracranial injections for successful delivery of X-gal to detect neurons that are highly-active after classical fear conditioning using PA imaging, now, we have further advanced this technique by using an intrathecal route of administration of X-gal combined with in vivo imaging to improve the translatability of this approach and whole-brain imaging capabilities. We subjected Fos/LacZ rats to fear conditioning, or control conditions. Ninety minutes post stimulus presentation, X-gal was delivered via intrathecal or intravenous injection. After 24 hours, a cranial window was made in animals and they were PA imaged in vivo. PA intensity within the medial prefrontal cortex (mPFC) of acquired images was quantified using ImageJ software. We presently report quantification of PA images of rat brains stained with X-gal product generated in vivo within activated neurons. Detailed PA signal differences are reported between mPFC subregions. We discuss the feasibility and translatability of this reporter method for neuronal activity based on our acquired images. With this technique, we propose a method of longitudinally monitoring activated neurons in vivo with high resolution and specificity.
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