Measurement of Blood Flow in the Area Postrema and Nucleus of the Solitary Tract Using <i>in vivo</i> 2-Photon Microscopy

The dorsal medulla is a major nervous system hub for autonomic regulation. This is due, in part, to two major nuclei: the area postrema (AP), a circumventricular organ, and the nucleus of the solitary tract (NTS). The AP is highly vascularized and contains fenestrated capillaries, allowing circulating factors to reach neurons responding to blood-borne signals. The NTS is an integrative center for visceral afferent information, which is then relayed to hypothalamic and limbic nuclei. Neurons in the NTS can also respond to circulating factors, including but not limited to glucose, leptin, GLP-1, and Angiotensin II. Thus, both regions have been intimately implicated in cardiovascular, body fluid, and metabolic regulation. Still, the anatomical and functional properties of the vascularization and blood flow within and between these dorsal brainstem regions, especially in response to shared physiological stimuli, remain largely unknown. This knowledge will help advance our understanding of how the brainstem senses and integrates circulating information to coordinate complex autonomic behaviors. To address this critical gap, we implemented a novel approach to measure blood flow in the dorsal brainstem using in vivo 2-photon (2P) microscopy in an anesthetized rat model. We have achieved this by creating a window through the dura and occipital bone, conserving the cerebellum, and exposing the AP and NTS. The local vasculature is then visualized via intravenous infusion of fluorescent dextrans, which allows for the identification and differentiation of arterioles, venules, and capillaries. To determine blood flow directionality and velocity quantitatively, 2P line scanning imaging and kymographs are used to assess red blood cell movement in these different vessels. To distinguish between the AP and NTS, microinjections of the fluorescent retrograde neuronal tracer Lumafluor are performed in the supraoptic and paraventricular nuclei of the hypothalamus, visualizing the somata of hypothalamic-projecting NTS neurons. With this novel approach, we have characterized the microcirculation properties of the AP and NTS for the first time in an in vivo preparation. Current experiments aim to assess neurovascular coupling responses following physiologically relevant homeostatic challenges. 1R01HL162575-01 This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.