Prediction of donor related lung injury in clinical lung transplantation using a validated ex vivo lung perfusion inflammation score.

BACKGROUND: Ex vivo lung perfusion (EVLP) is an isolated organ assessment technique that has revolutionized the field of lung transplantation and enabled a safe increase in the number of organs transplanted. The objective of this study was to develop a protein-based assay that would provide a precision medicine approach to lung injury assessment during EVLP. METHODS: Perfusate samples collected from clinical EVLP cases performed from 2009 to 2019 were separated into development (n = 281) and validation (n = 57) sets to derive and validate an inflammation score based on IL-6 and IL-8 protein levels in perfusate. The ability of an inflammation score to predict lungs suitable for transplantation and likely to produce excellent recipient outcomes (time on ventilator ≤ 3 days) was assessed. Inflammation scores were compared to conventional clinical EVLP assessment parameters and associated with outcomes, including primary graft dysfunction and patient care in the ICU. RESULTS: An inflammation score accurately predicted the decision to transplant (AUROC 68% [95% CI 62-74]) at the end of EVLP and those transplants associated with short ventilator times (AUROC 73% [95% CI 66-80]). The score identified lungs more likely to develop primary graft dysfunction at 72-hours post-transplant (OR 4.0, p = 0.03). A model comprised of the inflammation score and ∆PO2 was able to determine EVLP transplants that were likely to have excellent recipient outcomes, with an accuracy of 87% [95% CI 83-92]. CONCLUSIONS: The adoption of an inflammation score will improve accuracy of EVLP decision-making and increase confidence of surgical teams to determine lungs that are suitable for transplantation, thereby improving organ utilization rates and patient outcomes.

Activation of the Hypoglossal to Tongue Musculature Motor Pathway by Remote Control.

Reduced tongue muscle tone precipitates obstructive sleep apnea (OSA), and activation of the tongue musculature can lessen OSA. The hypoglossal motor nucleus (HMN) innervates the tongue muscles but there is no pharmacological agent currently able to selectively manipulate a channel (e.g., Kir2.4) that is highly restricted in its expression to cranial motor pools such as the HMN. To model the effect of manipulating such a restricted target, we introduced a "designer" receptor into the HMN and selectively modulated it with a "designer" drug. We used cre-dependent viral vectors (AAV8-hSyn-DIO-hM3Dq-mCherry) to transduce hypoglossal motoneurons of ChAT-Cre+ mice with hM3Dq (activating) receptors. We measured sleep and breathing in three conditions: (i) sham, (ii) after systemic administration of clozapine-N-oxide (CNO; 1 mg/kg) or (iii) vehicle. CNO activates hM3Dq receptors but is otherwise biologically inert. Systemic administration of CNO caused significant and sustained increases in tongue muscle activity in non-REM (261 ± 33% for 10 hrs) and REM sleep (217 ± 21% for 8 hrs), both P < 0.01 versus controls. Responses were specific and selective for the tongue with no effects on diaphragm or postural muscle activities, or sleep-wake states. These results support targeting a selective and restricted "druggable" target at the HMN (e.g., Kir2.4) to activate tongue motor activity during sleep.

GABA Cells in the Central Nucleus of the Amygdala Promote Cataplexy.

Cataplexy is a hallmark of narcolepsy characterized by the sudden uncontrollable onset of muscle weakness or paralysis during wakefulness. It can occur spontaneously, but is typically triggered by positive emotions such as laughter. Although cataplexy was identified >130 years ago, its neural mechanism remains unclear. Here, we show that a newly identified GABA circuit within the central nucleus of the amygdala (CeA) promotes cataplexy. We used behavioral, electrophysiological, immunohistochemical, and chemogenetic strategies to target and manipulate CeA activity selectively in narcoleptic (orexin-/-) mice to determine its functional role in controlling cataplexy. First, we show that chemogenetic activation of the entire CeA produces a marked increase in cataplexy attacks. Then, we show that GABA cells within the CeA are responsible for mediating this effect. To manipulate GABA cells specifically, we developed a new mouse line that enables genetic targeting of GABA cells in orexin-/- mice. We found that chemogenetic activation of GABA CeA cells triggered a 253% increase in the number of cataplexy attacks without affecting their duration, suggesting that GABA cells play a functional role in initiating but not maintaining cataplexy. We show that GABA cell activation only promotes cataplexy attacks associated with emotionally rewarding stimuli, not those occurring spontaneously. However, we found that chemogenetic inhibition of GABA CeA cells does not prevent cataplexy, suggesting these cells are not required for initiating cataplexy attacks. Our results indicate that the CeA promotes cataplexy onset and that emotionally rewarding stimuli may trigger cataplexy by activating GABA cells in the CeA.SIGNIFICANCE STATEMENT Although cataplexy has been closely linked to positive emotions for >130 years, the neural circuitry that underlies this relationship is poorly understood. Recent work suggests that the amygdala, a brain area important for processing emotion, may be part of this circuit. This study provides the first functional evidence to implicate GABA cells in the amygdala as regulators of cataplexy triggered by positive emotions and identifies the amygdala as the brain region important more for gating the entrance into rather than the exit from cataplexy. We also generated a new mouse model for studying GABA neurons in narcoleptic mice, which could serve as a useful tool for studying the neurobiological underpinnings of narcolepsy.

REM Sleep at its Core - Circuits, Neurotransmitters, and Pathophysiology.

Rapid eye movement (REM) sleep is generated and maintained by the interaction of a variety of neurotransmitter systems in the brainstem, forebrain, and hypothalamus. Within these circuits lies a core region that is active during REM sleep, known as the subcoeruleus nucleus (SubC) or sublaterodorsal nucleus. It is hypothesized that glutamatergic SubC neurons regulate REM sleep and its defining features such as muscle paralysis and cortical activation. REM sleep paralysis is initiated when glutamatergic SubC cells activate neurons in the ventral medial medulla, which causes release of GABA and glycine onto skeletal motoneurons. REM sleep timing is controlled by activity of GABAergic neurons in the ventrolateral periaqueductal gray and dorsal paragigantocellular reticular nucleus as well as melanin-concentrating hormone neurons in the hypothalamus and cholinergic cells in the laterodorsal and pedunculo-pontine tegmentum in the brainstem. Determining how these circuits interact with the SubC is important because breakdown in their communication is hypothesized to underlie narcolepsy/cataplexy and REM sleep behavior disorder (RBD). This review synthesizes our current understanding of mechanisms generating healthy REM sleep and how dysfunction of these circuits contributes to common REM sleep disorders such as cataplexy/narcolepsy and RBD.