Dysfunction of Astrocytes in the Nucleus Tractus Solitarii Leads to Cardiorespiratory Compromise

We have previously shown that saporin (SAP) microinjected into the nucleus tractus solitarii (NTS) causes loss of astrocytes while producing no immunohistochemical evidence of damage to NTS neurons. Loss of NTS astrocytes leads to loss of cardiovascular reflex control and, in some animals, sudden death as a result of asystole. Knowing that respiratory dysfunction and apnea may lead to asystole, we hypothesized that astrocytic dysfunction compromises respiratory control and may lead to apnea and sudden death. We sought to test that hypothesis in rats treated with bilateral NTS injections of the ribosomal toxin SAP, a chronic model of astrocytic loss, or fluorocitrate, which produces an acute model of astrocytic dysfunction. We first sought to determine if the loss of astrocytes due to SAP was due to necrosis or apoptosis by injection of SAP unilaterally into the NTS. We found loss of immunoreactivity for glial fibrillary acidic protein (GFAP), a marker of astrocytes, two hours, six hours, one day and seven days after SAP injection and increased immunoreactivity for the necrosis marker, receptor interacting protein (RIP), 6 hours and one day after SAP injection. In contrast, SAP treatment did not lead to apoptosis as documented by absence of positive cells in TUNEL reaction in the NTS. Injection of SAP into the NTS of neonatal rats (P13) led to loss of GFAP immunoreactivity seven days later as it also does in the adult rat brain. Therefore, in brain slices from P20–23 rats that had been treated unilaterally with SAP seven days previously, we performed patch clamp recording from neurons in the NTS and found that their intrinsic electrophysiological properties were unremarkable compared to neurons on the control side. In adult animals we monitored breathing chronically in a plethysmographic chamber after bilateral NTS microinjection of SAP. Seven days after injection, awake, treated rats, in contrast to control rats, began to manifest slow, irregular breathing patterns during eupneic breathing. During acute studies of astrocytic dysfunction, rats were anesthetized with isoflurane and respiration was monitored by intercostal electromyographic recording concurrent with recording arterial blood pressure and heart rate. Bilateral injections of fluorocitrate were made into NTS at 15 minute intervals for 2 hours. Fluorocitrate treated rats, but not rats treated with vehicle or PBS, developed slowed breathing followed by periodic apnea. These studies suggest that SAP causes necrosis of astrocytes and spares neuronal function. Either astrocytic necrosis or transient astrocytic dysfunction in the NTS interferes with normal cardiorespiratory regulation and leads to hypopnea and apnea that may contribute to death. Support or Funding Information NIH R01HL088090 and VA Merit Review