The central nervous system is critical in metabolic regulation, and accumulating evidence points to a distributed network of brain regions involved in energy homeostasis. This is accomplished, in part, by integrating peripheral and central metabolic information and subsequently modulating neuroendocrine outputs through the paraventricular and supraoptic nucleus of the hypothalamus. However, these hypothalamic nuclei are generally protected by a blood-brain-barrier limiting their ability to directly sense circulating metabolic signals—pointing to possible involvement of upstream brain nuclei. In this regard, sensory circumventricular organs (CVOs), brain sites traditionally recognized in thirst/fluid and cardiovascular regulation, are emerging as potential sites through which circulating metabolic substances influence neuroendocrine control. The sensory CVOs, including the subfornical organ, organum vasculosum of the lamina terminalis, and area postrema, are located outside the blood-brain-barrier, possess cellular machinery to sense the metabolic interior milieu, and establish complex neural networks to hypothalamic neuroendocrine nuclei. Here, evidence for a potential role of sensory CVO-hypothalamic neuroendocrine networks in energy homeostasis is presented.

Calcitonin gene-related peptide (CGRP) is a neuroactive peptide that is thought to play a role at efferent synapses in hair cell organs including the cochlea, lateral line, and semicircular canal. The deletion of CGRP in transgenic mice is associated with a significant reduction in suprathreshold cochlear nerve activity and vestibulo–ocular reflex (VOR) gain efficacy when compared to littermate controls. Here we asked whether the loss of CGRP also influences otolithic end organ function and contributes to balance impairments. Immunostaining for CGRP was absent in the otolithic end organs of αCGRP null (-/-) mice while choline acetyltransferase (ChAT) immunolabeling appeared unchanged suggesting the overall gross development of efferent innervation in otolithic organs was unaltered. Otolithic function was assessed by quantifying the thresholds, suprathreshold amplitudes, and latencies of vestibular sensory-evoked potentials (VsEPs) while general balance function was assessed using a modified rotarod assay. The loss of αCGRP in null (-/-) mice was associated with: (1) shorter VsEP latencies without a concomitant change in amplitude or thresholds, and (2) deficits in the rotarod balance assay. Our findings show that CGRP loss results in faster otolith afferent activation timing, suggesting that the CGRP component of the efferent vestibular system (EVS) also plays a role in otolithic organ dynamics, which when coupled with reduced VOR gain efficacy, impairs balance.

Hypertension affects nearly half of US adults and is a leading cause for heart attack and stroke. Angiotensin II (Ang-II) is a well-recognized driver of hypertension, particularly through its sympathoexcitatory actions within the central nervous system (CNS). As a peptide hormone, Ang-II is too large to cross into the brain and acts at circumventricular regions lacking a blood-brain-barrier, particularly the forebrain subfornical organ (SFO). While Ang-II has been shown to drive pro-hypertensive cellular stressors in the SFO (i.e. oxidative and endoplasmic reticulum stress), it is unclear how these mechanisms translate into long-term alterations at the cellular level. Intriguingly, stress-associated pathways can induce cellular senescence and the senescence-associated secretory phenotype (SASP). Chronic senescence/SASP leads to detrimental changes in cell metabolism, macromolecule damage, and a pro-inflammatory environment capable of propagating senescence. We previously demonstrated cellular senescence at the mRNA level, including robust upregulation of the key senescence mediator p16 ( CDKN2A), in the SFO during Ang-II-induced hypertension. However, the cell type(s) that undergoes senescence in response to Ang-II remains unknown. Thus, we hypothesized, that Ang-II would elicit cellular senescence at the protein level and furthermore that this would occur in both SFO neurons and astrocytes. To test this, male C57Bl/6J mice were implanted with subcutaneous osmotic minipumps for chronic infusion of Ang II (600 ng/kg/min) for 0 or 14 days (n=4-5/group), and brains were collected and prepared for immunohistochemistry (IHC). Immunohistochemical analysis revealed a marked ~46% increase in p16 intensity in the SFO following 14 days of Ang-II infusion (Integrated density: 1.00±0.08 vs 1.46±0.24, Day 0 vs Day 14, p=0.07). The increase in p16 integrated density occurred throughout rostral to caudal SFO, most notably in the medial region. We next sought to characterize the SFO cell types that undergo cellular senescence using double immunohistochemistry for p16 and either neurons (NeuN) or astrocytes (GFAP). Following 14 days of Ang-II administration and histological procedures, colocalization analysis was performed using IMARIS software. When first examining neuronal populations, extremely low to no co-localization with p16 was found (p16/NeuN: Pearson’s correlation coefficient=-0.074, Mander’s overlap coefficient=0.071). However, p16 was very highly co-localized with astrocytes (p16/GFAP: Pearson’s correlation coefficient=0.545, Mander’s overlap coefficient=0.992). Together, these findings indicate that: 1) p16-associated cellular senescence occurs at the protein level in the SFO during Ang-II-induced hypertension; and 2) p16 is primarily upregulated in SFO astrocytes, but not neurons, during hypertension development. R01DK117007, R01HL141393, F31HL164059 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Hypertension affects 1 in 3 US adults and is a leading risk factor for heart attack and stroke. The peptide hormone angiotensin II (Ang II) is a well‐recognized driver of hypertension, particularly through its sympathoexcitatory actions within the central nervous system (CNS). Although a number of pro‐hypertensive CNS mechanisms (i.e. neurogenic hypertension) have been elucidated, including oxidative and endoplasmic reticulum stress, how these mechanisms translate into long‐term alterations in CNS circuits remains unclear. Intriguingly, stress‐associated pathways can culminate in cellular senescence and the senescence‐associated secretory phenotype (SASP). Chronic senescence/SASP leads to marked changes in cell metabolism, macromolecule damage, and a pro‐inflammatory environment. Based on this, we hypothesized that CNS cellular senescence may be a key contributor to neurogenic hypertension. We first performed a proof‐of‐principle experiment to determine if CNS cellular senescence is involved in blood pressure regulation. C57Bl/6J male mice were fitted with intracerebroventricular (ICV) cannulas and underwent implantation of radiotelemeters for conscious recording of cardiovascular parameters. Following surgical recovery, the senescence inducing agent doxorubicin (0.00125 mg) or vehicle control was administered daily over three days (n=3/group). Daily ICV administration of doxorubicin resulted in marked elevations in mean arterial blood pressure within 48 hours that were sustained throughout the study (72 hours: 107±1 vs. 123±1 mmHg; ICV vehicle vs. doxorubicin, p<0.05). Relative to controls, ganglionic blockade (i.p. chlorisondamine 12 mg/kg) elicited a greater fall in blood pressure in doxorubicin‐treated animals (Δ‐11±4 vs. Δ‐56±6 mmHg; ICV vehicle vs. doxorubicin, p<0.05). Given these findings that CNS cellular senescence is associated with a hypertensive phenotype, likely through alterations in autonomic control of blood pressure, we next profiled cellular senescence in key cardioregulatory nuclei during hypertension development. Male C57Bl/6J mice were implanted with subcutaneous osmotic minipumps for chronic infusion of Ang II (600 ng/kg/min). Brains were collected at baseline and after 14 days of Ang II infusion (n=4‐5/group) and micropunches of cardiovascular and autonomic nuclei including the organum vasculosum lamina terminalis (OVLT), subfornical organ (SFO), and paraventricular nucleus of the hypothalamus (PVN) were collected. Two‐week infusion of Ang II resulted in a robust increase in the key senescent gene p16 (CDKN2A) in the SFO (6.1±0.8 fold baseline, p<0.05). Interestingly, Ang II‐induced hypertension was not associated with changes in p16 in the OVLT (2.2±0.7 fold baseline, p=0.2) and PVN (1.5±0.4 fold baseline, p=0.4). Additionally, Ang II‐induced senescence in the SFO was paralleled by the upregulation of SASP indicators (e.g. Interleukin‐1α: 6.5±1.4 fold baseline, p<0.05). Together, these findings indicate that: 1) CNS cellular senescence is pro‐hypertensive; and 2) Ang II elicits cellular s...

Sex differences in the development of hypertension across the lifespan are well established. Women are at lower risk of hypertension compared to age‐matched men until menopause, when the risk of hypertension sharply rises for women. The peptide hormone angiotensin II (Ang II) is a well‐recognized contributor to hypertension, particularly through its actions in the brain. Importantly, Ang II is too large to cross into the central nervous system (CNS) and influences blood pressure regulation by acting at circumventricular nuclei located outside of the blood‐brain‐barrier, particularly the forebrain subfornical organ. For instance, in males, Ang II drives pro‐hypertensive cellular stressors in the SFO while females are resistant to Ang II‐induced hypertension. However, the underlying CNS (i.e. SFO) mechanisms that contribute to hypertension in a sexually dimorphic manner remain unclear. Intriguingly, Ang II‐induced stressors can culminate in cellular senescence. Cellular senescence is a complex cellular phenotype characterized by marked changes in cell metabolism, macromolecular damage, and a pro‐inflammatory environment known as the senescence associated secretory phenotype (SASP). The role of senescence in the CNS as related to the development of hypertension remains unclear. Based on this, we hypothesized that brain cellular senescence, particularly in the SFO, may be a novel mechanism for the sexually dimorphic nature of Ang II‐induced hypertension. To test this, 8‐week‐old male and female C57Bl/6J mice (n=7‐10/group) were implanted with subcutaneous mini‐osmotic pumps for the infusion of Ang II (600 ng/kg/min). Micropunches of the SFO were collected at baseline and following 14 days of Ang II infusion for quantitative real‐time PCR analysis. In males, infusion of Ang II resulted in a robust increase in key senescent genes including p16 (1.9±0.3 fold baseline, p<0.05) and p21 (2.5±0.5 fold baseline, p<0.05) in the SFO. In parallel with the induction of senescence, inflammatory SASP indicators were also upregulated in the SFO of males following 14 days of Ang II infusion, including interleukin‐6 (IL‐6: 2.4±0.4 fold baseline, p<0.05) and interleukin‐10 (IL‐10: 2.8±0.7 fold baseline, p<0.05). However, when examining females, clear sexual dimorphism in Ang II‐induced SFO cellular senescence was apparent. Specifically, no changes in p16 (1.1±0.4 fold baseline, p=0.9) or p21(1.2±0.2 fold baseline, p=0.2) were noted in the SFO following Ang II administration. Similarly, SASP markers that accompany senescence remained unchanged in the SFO (e.g. IL‐6: 1.3±0.5 fold baseline, p=0.8 and IL‐10: 0.98±0.2 fold baseline, p=0.6). Together, these findings indicate that: 1) Cellular senescence in the SFO is associated with Ang II‐induced hypertension in males; and 2) Females are protected against Ang II‐driven cellular senescence in the SFO. Importantly, cellular senescence increases with advancing age. Thus, our data suggests that cellular senescence may underlie the sexually dimorphic nature of hypertension at younger ages, as w...