Integrated Ultrasound Neuromodulation and Optical Neuroimaging in Awake Mice using a Transparent Ultrasound Transducer Cranial Window

Ultrasound neuromodulation is a rapidly advancing, non-invasive technique with significant therapeutic potential for treating various neurological disorders. Although extensive in vitro and in vivo studies have provided valuable insights into its modulatory effects, the underlying mechanisms remain poorly understood, limiting its clinical translation. Optical neuroimaging techniques can help investigate these mechanisms; however, the opacity and bulkiness of conventional ultrasound transducers pose significant challenges for their integration with in vivo ultrasound neuromodulation studies, particularly in awake rodents. To address these limitations, we propose a straightforward solution: a miniaturized lithium niobate-based transparent ultrasound transducer (TUT) integrated as a thinned-skull cranial window for ultrasound stimulation while facilitating multimodal optical neuroimaging in awake mice brain. Using laser speckle contrast imaging and intrinsic optical signal imaging, we studied changes in brain hemodynamics in response to various ultrasound stimulation sequences. Our experiments demonstrated that TUT cranial window can robustly induce neuromodulatory effects with observed increase in both cerebral blood flow and total hemoglobin, with peak and cumulative hemodynamic changes directionally correlated with ultrasound stimulation duration and intensity. Overall, these findings highlight that TUT cranial window can seamlessly integrate ultrasound stimulation and optical neuroimaging in awake mouse brain models, offering promising prospects for uncovering the underlying mechanisms of ultrasound neuromodulation.