Limbic system involvement in modulation of breathing during seizures and arousal

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Automatic control of breathing is mediated by a neural network in the brainstem, modulated by central and peripheral inputs. A major source of this modulation comes from chemoreceptors from peripheral and central level. In addition, automatic control of breathing can be overridden by voluntary or conscious control. Pathways involved in this conscious control may be usurped by seizures, resulting in ictal apnea, at the same time that chemoreceptor drive is impaired. Here we studied mechanisms by which the limbic system might modulate breathing, and how this may lead to fatal apnea induced by a seizure. We used Scn1aR1407X/+ mice, a genetic mouse model of Dravet Syndrome (DS) and SUDEP, to study how activation of the Central Amygdala (CeA) can modify breathing and induce fatal apneas. Methods In DS mice (n=4), the CeA was stimulated under light anesthesia by injecting current with monopolar electrodes guided by a stereotaxic manipulator while measuring breathing using head-out plethysmography. The locations of the electrode were verified post hoc by electrolytic lesion and histology. A 3D map was generated of the portions of the CeA where breathing was modulated. In a separate group of DS (n=7) mice, whole-body plethysmography was used to assess baseline breathing, the hypercapnic ventilatory response (HCVR) and the hypoxic ventilatory response (HVR) before and after lesioning the CeA. In a third group of DS mice (n=11), electrolytic lesions were induced in the CeA. Mice were then instrumented with headmounts with EEG, ECG and EMG electrodes. After 5 days of recovery, headmounts were connected to a preamplifier and a commutator, and mice were placed in a plethysmography chamber to monitor breathing as well as EEG, ECG and EMG activity. Body temperature was increased with a heat lamp by 0.5°C per minute until a seizure was induced, or a maximum of 42.5°C was reached. Results CeA lesions did not affect baseline breathing, the HCVR or HVR, and did not prevent seizures in DS mice. Respiratory arrest and death occurred in control (sham-lesioned) DS mice following heat-induced seizures in 54.4% of cases (n=11). In contrast, only 9.1% of DS mice with CeA lesions showed ventilatory arrest and death after heat-induced seizures (n=11; p=0.016). Unilateral rostral amygdala lesions increased the survival rate from spontaneous seizures in DS mice (26.10% vs 49.12.5%, n=14 control, n=20 lesioned p=0.022). Stimulation of the rostral CeA produced apnea without causing seizures. Complete apnea could be induced for 1 minute without causing death in three animals, and for four minutes in one animal. Breathing returned immediately upon termination of amygdala stimulation in each case. There was also no reversal of anesthetic immobility, suggesting lack of an arousal response. Conclusions Our data suggest that the CeA modulates breathing and can induce complete apnea but is not directly involved in baseline respiratory rhythm generation or the ventilatory response to hypercapnia or hypoxia. Lesions of the CeA reduce the risk of apnea during seizures. We conclude that the CeA is a critical component of the neural pathway responsible for spread of seizures in the forebrain to key respiratory centers in the brainstem that elicits ventilatory arrest.