Fogarty, Matthew J.

In mammals, breathing requires an inspiratory pump to generate a negative intrathoracic pressure and thus pull air into the lungs for gas exchange. Exclusively in mammals, the diaphragm muscle (DIAm) is the major inspiratory pump, which separates the thoracic and abdominal cavities. With DIAm contraction, a negative intrathoracic and positive abdominal pressure are generated [i.e., transdiaphragmatic pressure (P di )]. During breathing, the DIAm is very active, with a duty cycle (time active vs. inactive) similar to that of the heart. Like the heart, this is a pump that cannot fail! Thus, in controlling breathing, the nervous system must activate DIAm to accomplish pump function while avoiding fatigue. The timing of DIAm activations must also be coordinated with activation of the upper airway muscles (the pipes) to avoid airway occlusion and aspiration and the intercostal and abdominal muscles to appropriately stiffen the body cavities. Similarly, neural control and coordination of the DIAm in nonventilatory behaviors (airway protection, swallowing, vocalization, and voiding) are equally complex, with coordinated activation of the DIAm and abdominal muscles. It is becoming increasingly clear that impaired nonventilatory functions underlie the pathophysiology of a variety of medical conditions. In this review, we will delve into the detailed mechanistic underpinnings of the neural control of the DIAm and the symphonic coordination of its activation during breathing and other behaviors. We will attempt to move the field from considering the DIAm only as an inspiratory pump, emphasizing neural control of airway, intercostal, and abdominal muscles that must be coordinated with DIAm activation.

Age-associated muscle weakness and atrophy (sarcopenia) is prevalent in both limb and respiratory muscles. In the limb muscles, dyscoordination and sarcopenia predisposes elderly individuals to a greater risk of fall hospitalization, protracted recovery and subsequent frailty. In humans and in the Fischer 344 rat model of aging, motor neuron (MN) death occurs in old age, and the selective vulnerability of fast-fatigable motor units to loss is consistent with behavioral deficits. In many behaviors, deficits in neuromotor coordination and the timing of motor unit activations may be more pertinent than frank weakness. In MNs, the substrate for the integration of sensory, pattern, executive and reflex activations is the dendritic arbour. Currently, it is unknown whether MN dendritic and dendritic spine degenerations occur in aging and if any changes are specific to the vulnerable larger MNs. In this study of female and male Fischer 344 rats at 6- and 24-months-old, we evaluated lumbar MN dendritic arbours and dendritic spines using golgi-cox impregnation and confocal imaging approaches in thick horizontal vibratome ex vivo slices (200 µm). All data were analyzed by unpaired t-tests, 2-way ANOVAs or Pearson’s correlations, where relevant. In 24-month-old rats, lumbar MN dendritic arbour length and surface area were reduced compared to 6-month controls. In 6-month-olds, dendritic length and surface area was dependent on somal size, with bigger MN somas having longer and larger dendritic arbours, a relationship lost in 24-month old lumbar MNs. Dendritic spine density was reduced in 24- compared to 6-month-old rats. Overall, our findings suggest that lumbar MN dendritic degenerations with aging are selective to MNs of fast-fatiguable motor units. Interventions ameliorating neuromotor circuit and dendritic breakdown may preserve neuromotor function in old age. This work was supported by NIH grants: HL166204 & AG086136 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.

Phrenic motor neurons (PhMNs) innervate the diaphragm muscle (DIAm), the primary pump muscle involved in breathing. It has been shown that brain-derived neurotrophic factor (BDNF) signals through its high-affinity, full-length tropomyosin receptor kinase B (TrkB.FL) to promote PhMN survival in embryonic and early postnatal development. Reduced BDNF/TrkB.FL signaling also appears to be involved in the age-related loss of PhMNs. Previously, we developed a novel chemogenetic rat model ( TrkB F616 rat) in which insertion of a knock-in allele sensitive to 1NMPP1 (a selective kinase inhibitor) allows rapid inhibition of TrkB.FL kinase activity. Using this TrkB F616 rat, we found that in younger animals, a 14-day period of 1NMPP1-induced TrkB.FL kinase inhibition resulted in an ~20% loss of larger PhMNs similar to that found with aging. Previous studies from our laboratory demonstrated that BDNF/TrkB.FL signaling induces pCREB S133 phosphorylation at serine residue 133 in PhMNs. 1NMPP1 induced inhibition of TrkB.FL kinase activity in TrkB F616 rats acutely blunts pCREB S133 phosphorylation induced by intrathecal BDNF treatment. In another cell type, we showed that pCREB S133 transcriptionally targets PGC1α expression with downstream upregulation of expression of mitochondrial transcription factor A (TFAM) and mitochondrial biogenesis. Based on a bioinformatic analysis, we identified a putative binding site for pCREB S133 in the rat PGC1α promoter. In the present study, we hypothesized that BDNF/TrkB.FL signaling mediates pCREB S133 phosphorylation and downstream activation of PGC1α thereby promoting mitochondrial biogenesis. Adult female and male TrkB F616 rats (~ 3 months of age; 300 g) were anesthetized (ketamine/xylazine) and either 25 μM 1NMPP1 or DMSO (vehicle control) was injected ip. After 2 h, the rats were treated intrathecally (infused at C 4 ) with 100 ng/ml BDNF for 40 min. Thereafter, the animals were euthanized and the ventral cervical spinal cord containing PhMNs was dissected, and lysates were prepared for subsequent analyses. Western blot was performed to quantify pCREB S133 phosphorylation, and PGC1α mRNA and protein expression were measured by qPCR and Western blot, respectively. A chromatin immunoprecipitation (ChIP) assay confirmed that pCREB S133 binds to the PGC1α promoter and that compared to vehicle treated controls, 1NMPP1induced TrkB.FL kinase inhibition reduced the binding affinity of pCREB S133 . Consistent with these results, we found that pCREB S133 phosphorylation and PGC1α mRNA and protein expression were reduced following 1NMPP1-induced TrkB.FL kinase inhibition. We conclude that in the rat cervical ventral horn, BDNF/TrkB.FL signaling induces pCREB S133 phosphorylation which transcriptionally activates PGC1α expression and downstream mitochondrial biogenesis. The BDNF/TrkB.FL signaling pathway acting in concert with activity dependent signaling may converge on pCREB S133 phosphorylation to maintain the survival of PhMNs by promoting mitochondrial biogenesis. The loss of the contribution of BDNF/TrkB.FL signaling in old age and with neurodegeneration may lead to PhMN death. Supported by NIH grants AG44615 and HL146114 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.

In the elderly, age-associated muscle weakness and atrophy (sarcopenia) is prevalent in both limb and respiratory muscles. In the tongue muscles, dyscoordination and sarcopenia predisposes elderly individuals to dysphagia, dysphonia and a greater risk for aspiration pneumonias and obstructive sleep apnea. In humans and in the Fischer 344 rat model of aging, motor neuron (MN) death occurs in old age, and the selective vulnerability of fast-fatigable motor units to loss is consistent with behavioral deficits in airway defense (e.g., cough). However, whether the timing of MN loss is gradual across the lifespan or precipitous from late-middle-age is unknown. Recently, the concept of an age -related increase of systemic inflammatory cytokines has been implicated in the development of sarcopenia, although the mechanisms by which this occurs remain obscure. In other cell types, inflammation triggers an unfolded protein response and mitochondrial fragmentation – processes implicated in neuronal death and neurodegeneration. In this study of female and male Fischer 344 rats at 6-, 18- and 24-months-old, we evaluated serum inflammatory cytokines (TNFα, IL1β & IL6) via ELISA, medullary unfolded protein response (pIRE1a S724 & CDK5) and mitochondrial fragmentation (pDRP1 S616 ) via Western blot, hypoglossal MN mitochondrial volume density and mitochondrial fragmentation via serial blockface electron microscopy (SBFEM), hypoglossal MN survival via Nissl histology and stereological counting and tongue muscle (longitudinal and genioglossus) sarcopenia via uniaxial specific force assessment and cross-sectional muscle fiber areas. All data were analyzed by 1-way ANOVAs or Kruskal-Wallis nonparametric tests, depending on data distributions. Here, we show that a systemic inflammation-induced unfolded protein response in the medulla and hypoglossal MNs mitochondrial fragmentation is present at 18-months-old. This mitochondrial fragmentation is consistent with increased medullary expression of pDRP1 S616 . At this age, we did not observe any evidence of tongue muscle weakness or atrophy, nor any evidence of hypoglossal MN death. At 24-months old, hypoglossal MN mitochondrial deficits remained readily observable using SBFEM, with hypoglossal MN death (specific to larger MNs) and tongue muscle sarcopenia (selective to fast-fatigable type IIx fibers) present at this age. Overall, we conclude that denervation-related sarcopenia is precipitous from late-middle-age to old age in Fischer 344 rats. This MN death is triggered (or at least preceded), by an unfolded protein response and mitochondrial fragmentation. Interventions ameliorating inflammation, the unfolded protein response and/or mitochondrial breakdown may preserve MNs and prevent or reduce age-associated sarcopenia. This work was supported by NIH grants: HL166204 (MJF), AG086136 (MJF), HL157984 (GCS) & AG44615 (GCS) 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.

Age-associated muscle weakness and atrophy (sarcopenia) is prevalent in both limb and respiratory muscles. In the tongue muscles, dyscoordination and sarcopenia predisposes elderly individuals to dysphagia, dysphonia and a greater risk for aspiration pneumonias and obstructive sleep apnea. In humans and in the Fischer 344 rat model of aging, motor neuron (MN) death occurs in old age, and the selective vulnerability of fast-fatigable motor units to loss is consistent with behavioral deficits in airway defense (e.g., cough). However, in many aerodigestive behaviors, deficits in neuromotor coordination and the timing of motor unit activations may be more pertinent than frank weakness. In motor neurons, the substrate for the integration of sensory, pattern, executive and reflex activations is the dendritic arbour. Currently, it is unknown whether MN dendritic and dendritic spine degenerations occur in aging and if any changes are specific to the vulnerable larger MNs. In this study of female and male Fischer 344 rats at 6- and 24-months-old, we evaluated hypoglossal MN dendritic arbours and dendritic spines using golgi-cox impregnation and confocal imaging approaches in thick brainstem vibratome ex vivo slices (200 µm). All data were analyzed by unpaired t-tests, 2-way ANOVAs or Pearson’s correlations, where relevant. In 24-month-old rats, hypoglossal MN dendritic arbour length and surface area were reduced compared to 6-month controls. In 6-month-olds, dendritic length and surface area was dependent on somal size, with bigger MN somas having longer and larger dendritic arbours, a relationship lost in 24-month old hypoglossal MNs. Dendritic spine density was reduced in 24- compared to 6-month-old rats. Overall, our findings suggest that hypoglossal MN dendritic degenerations with aging are selective to MNs of fast-fatigable motor units. Interventions ameliorating neuromotor circuit and dendritic breakdown may preserve neuromotor function in old age. This work was supported by NIH grants: HL166204 & AG086136 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.

Neuromotor control of diaphragm muscle (DIAm) involves size-dependent recruitment of phrenic motor neurons (PhMNs). To accomplish breathing, only slow (type S) and fast, fatigue resistant (type FR) DIAm motor units are recruited. Thus, type s and FR DIAm motor units are very active in sustaining ventilation throughout life. Importantly, type S and FR DIAm motor units comprise small PhMNs that innervate type I and IIa DIAm fibers, respectively. More fatigable fast (type FF) motor units are very infrequently recruited for higher force, expulsive behaviors. Type FF DIAm motor units comprise large PhMNs that innervate type IIx/IIb DIAm fibers. Previously, we reported that compared to large PhMNs, small PhMNs have higher mitochondrial volume densities reflecting their more frequent activation. Using a quantitative histochemical technique to measure the maximum velocity of the succinate dehydrogenase reaction (SDH max ), we showed that type I and IIa DIAm fibers have higher oxidative capacity compared to type IIx/IIb fibers, which correlates with mitochondrial volume density and the more frequent activation of these muscle fibers. Accordingly, we hypothesize that the SDH max of small PhMNs is greater compared to large PhMNs. PhMNs in 6 month old, female and male Sprague Dawley rats were retrogradely identified by intrapleural injection of Alexa 647-conjugated cholera toxin B (CTB). Following retrograde CTB labeling, rats were perfused intracardially with a 0.1 M PBS, euthanized, and the C1–C6 segment of the spinal cord was dissected. Spinal cord sections were cut (40 mm) in longitudinal horizontal sections and mounted in 2-well Ibidi μ-slide plates. To measure SDH max , the sections were exposed to a solution containing 80 mM succinate (maximum substrate concentration) and 1.5 mM nitro blue tetrazolium (NBT; reaction indicator). As the SDH reaction proceeded, CTB-labeled PhMNs were imaged in 3D (Z optical slice of 0.5 μm) using an oil-immersion x60/1.4 NA objective on a Nikon Eclipse A1 laser scanning confocal system. The change in optical density (OD) was measured every 15 s over a 10-min period to measure the rate of accumulation of NBT diformazan (NBT dfz ; peak absorbance of 570 nm) in PhMNs. Using the Beer-Lambert equation, the change in OD was used to determine SDH max expressed as mmol fumarate/L cell/min. In separate experiments, following retrograde CTB labeling, mitochondria in PhMNs were labeled by transdural infusion of MitoTracker Red and imaged in 3D to determine mitochondrial volume density. Our results support a size-dependent heterogeneity in PhMN mitochondrial volume density and SDH max . We conclude that more active small PhMNs have a higher mitochondrial volume density and corresponding SDH max compared to large PhMNs. NIH grant AG44615 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.

Histamine appears to be intimately involved with skeletal muscle responses during and following exercise. Blocking histamine’s actions during endurance exercise training by taking common over-the-counter antihistamines resulted in diminished gains in fitness. These results indicate that exercise-induced histamine release is important in generating many of the positive adaptations to exercise training.

ABSTRACT Symmorphosis is a concept of economy of biological design, whereby structural properties are matched to functional demands. According to symmorphosis, biological structures are never over designed to exceed functional demands. Based on this concept, the evolution of the diaphragm muscle (DIAm) in mammals is a tale of two structures, a membrane that separates and partitions the primitive coelomic cavity into separate abdominal and thoracic cavities and a muscle that serves as a pump to generate intra‐abdominal (P ab ) and intrathoracic (P th ) pressures. The DIAm partition evolved in reptiles from folds of the pleural and peritoneal membranes that was driven by the biological advantage of separating organs in the larger coelomic cavity into separate thoracic and abdominal cavities, especially with the evolution of aspiration breathing. The DIAm pump evolved from the advantage afforded by more effective generation of both a negative P th for ventilation of the lungs and a positive P ab for venous return of blood to the heart and expulsive behaviors such as airway clearance, defecation, micturition, and child birth. © 2019 American Physiological Society. Compr Physiol 9:715‐766, 2019.

Abstract The mammalian neuromuscular junction (NMJ) comprises a presynaptic terminal, a postsynaptic receptor region on the muscle fiber (endplate), and the perisynaptic (terminal) Schwann cell. As with any synapse, the purpose of the NMJ is to transmit signals from the nervous system to muscle fibers. This neural control of muscle fibers is organized as motor units, which display distinct structural and functional phenotypes including differences in pre‐ and postsynaptic elements of NMJs. Motor units vary considerably in the frequency of their activation (both motor neuron discharge rate and duration/duty cycle), force generation, and susceptibility to fatigue. For earlier and more frequently recruited motor units, the structure and function of the activated NMJs must have high fidelity to ensure consistent activation and continued contractile response to sustain vital motor behaviors (e.g., breathing and postural balance). Similarly, for higher force less frequent behaviors (e.g., coughing and jumping), the structure and function of recruited NMJs must ensure short‐term reliable activation but not activation sustained for a prolonged period in which fatigue may occur. The NMJ is highly plastic, changing structurally and functionally throughout the life span from embryonic development to old age. The NMJ also changes under pathological conditions including acute and chronic disease. Such neuroplasticity often varies across motor unit types. © 2022 American Physiological Society. Compr Physiol 12: 3731–3766, 2022.