Citations

Abstract Background Heart failure (HF) is classified by ejection fraction: reduced EF (<40%) is HFrEF and preserved EF (>50%) is HFpEF. Unlike HFrEF, no therapeutic agent improves mortality in HFpEF. The molecular mechanism that produces HFpEF is not completely understood, but the cascade of pathology that produces HFpEF is thought to begin with changes in vascular reactivity, including a decrease in NO mediated vasodilatation, which coupled with subsequent changes in contractility, energetics and coronary blood flow produce HFpEF. If abnormal vascular reactivity is the initial step in the pathological cascade that produces HFpEF, restoring and/or improving vascular reactivity could represent a novel treatment strategy. Vascular reactivity is primarily regulated by myosin light chain phosphatase, which has catalytic, myosin targeting (MYPT1) and 20kDa subunits. Alternative mRNA splicing of exon24 (E24) of the MYPT1 transcript produces MYPT1 isoforms that differ by the presence or absence of a COOH-terminal leucine zipper (LZ+/LZ-); E24 exclusion produces an NO responsive LZ+ MYPT1, while E24 inclusion produces an NO unresponsive LZ- MYPT. Methods We used the mouse two-hit model of HFpEF (high fat diet and L-NAME) and treated mice with an anti-sense octo-guanidine targeting the 5’ splice site of E24 (ASO-E24) to increase the expression of the NO responsive, LZ+ MYPT1 isoform in vascular smooth muscle. Invasive and noninvasive hemodynamics were used to determine LV function. Results Compared to mice with HFpEF, ASO-E24 treatment maintains LZ+ MYPT1 expression (4.7±0.7au v 1.0±0.4au v 2.0±0.4au, control v HFpEF v ASO-E24 Rx, p<0.05), improves diastolic function; LVEDP (10±1mmHg v 20±4mmHg v 14±3mmHg, p<0.05), dP/dtmin (−8000±300mmHg/s v 6000±500mmHg/s v 8500±700mmHg/s, p<0.05), both early (E; 0.60±0.05m/s v 0.42±0.06m/s v 0.64±0.06m/s, p<0.05) and late diastolic filling (A; 0.38±0.03m/s v 0.24±0.02m/s v 0.47±0.04m/s, p<0.050 and also prevents the increase in lung weight (167±5g v 175±7g v 166±5g, p<0.05). Further, mice treated with ASO-E24 maintained normal relaxation to 8Br-cGMP (65±5% v 44±9% v 72±9%, p=0.05). Conclusion These data demonstrate that maintaining normal LZ+ MYPT1 expression and vascular reactivity prevent the development of HFpEF. These results are consistent with the hypothesis that abnormal vascular reactivity is the initial and primary step in the pathological cascade that produces HFpEF and ASO-E24, which is designed to preserve normal LZ+ MYPT1 expression and vascular reactivity, could represent a novel and effective treatment strategy for HFpEF. Clinical Prospective What is New? In HFpEF, an anti-sense octo-guanidine targeting the 5’ splice site of exon 24 (ASO-E24) of the MYPT1 transcript increases the expression of the NO responsive, LZ+ MYPT1 isoform in vascular smooth muscle. Preserving LZ+ MYPT1 expression and normal vascular reactivity prevents the development of HFpEF. A decrease in LZ+ MYPT1 expression contributes to the development of HFpEF. What are the Clinical Implications? The results of the study demonstrate that abnormal vascular reactivity is an important step in the pathological cascade that produces HFpEF Increasing LZ+ MYPT1 expression could represent a novel treatment strategy for HFpEF.

Abstract Striated muscle weakness and motor neuron (MN) death are associated with aging. These alterations affect both brainstem and spinal motor pools, including hypoglossal MNs innervating the tongue and lumbar MNs innervating the lower/hindlimbs. Both MNs and striated muscles exhibit type‐selective motor unit vulnerability, where larger MNs (likely fast fatigable motor units) and type IIx/b muscles are vulnerable to death and atrophy, respectively. These findings have been cemented in humans to some degree and in Fischer 344 and Wistar rats. However, MN survival in old age has not been evaluated in Sprague–Dawley rats. Here, in 6‐ (young) and 24‐month‐old (old) female and male Sprague–Dawley rats, we performed stereological MN counts on hypoglossal and lumbar MNs using 16 μm Nissl‐stained sections. We observed a 35% loss of hypoglossal and lumbar MNs in old age. Although mean MN somal surface areas were moderately reduced in hypoglossal and unchanged in lumbar MNs with age, frequency histogram analyses revealed a disproportionate loss of larger MN in old age hypoglossal and lumbar pools. These findings reveal reliable and robust observations of MN death in rats. This neurogenic aspect of neuromotor impairment with age may be a key driver of sarcopenia and frailty in the elderly.

Abstract Age‐associated muscle weakness and atrophy of limb muscles, termed sarcopenia, is a major factor in the morbidity of the elderly. It is becoming increasingly recognized that a substantial contribution to the sarcopenic phenotype arises from motor neuron (MN) death in old age and subsequent muscle denervation. In human and rodent models, mitochondrial dysfunction is a leading culprit in both muscle and MN deterioration with age. In other motor pools within the Fischer 344 (F344) aging rat model, we showed that sarcopenia is selective to type IIx/b muscle fibers. We also showed a loss of larger MNs and denervation of the IIx/b fibers, which together comprise more fatigable fast (type FF) motor units. This selective vulnerability of larger MNs and type IIx/b muscle fibers to sarcopenia is further reflected by reductions in oxidative capacity of IIx/b fibers, as assayed by determining the maximum velocity of the succinate dehydrogenase reaction (SDHmax). Here, we hypothesize similar changes will occur in F344 tibialis anterior (TA) muscle from young (6‐months) and old (24‐months) groups. We also predict lumbar MN death in young and old F344 rats. We found selective atrophy of type IIx/b fibers in the TA from old rats. In old age, SDHmax was reduced but only in IIx/b TA fibers. These muscle observations were concomitant with the death of larger lumbar MNs in old age. This study indicates the remarkable selectivity of type FF TA motor units to age‐associated perturbations and sarcopenia.

Aging selectively impacts the function of type IIx/IIb fibers in the diaphragm muscle. We show that sarcopenia reduces maximum specific force, maximum shortening velocity, and peak power output, whereas endurance during repetitive contractions is preserved. These findings explain why higher force airway clearance behaviors decline with age, whereas fatigue-resistant breathing behavior remains resilient.

Neuroplastic changes in phrenic motor neurons that underlie recovery from spinal cord injury may be mediated by perineuronal nets. We show that changes in perineuronal nets around phrenic motor neurons occur within 7 days after cervical spinal cord injury, and that the increase in perineuronal nets around larger phrenic motor neurons was more pronounced. These changes are consistent with substantial reorganization of the neural network onto intact phrenic motor neurons following cervical spinal cord injury.

Abstract Age‐related neuromuscular dysfunction is widely recognized; however, the extent to which aging is accompanied by motor neuron (MN) loss remains underexplored. This review evaluates evidence for MN loss with aging across species and anatomical motor pools. We synthesize PUBMED studies in humans, non‐human primates, rats and mice within the spinal cord and brainstem. Attention was given to the methodological approach used to quantify MNs and categorized based on what MNs were quantified (specific pools, vertebral levels, lateral motor columns). Methods were assessed qualitatively, based on stereological and non‐stereological density counting approaches, and the staining techniques identified as optimal or suboptimal by defined criteria. To facilitate cross‐study comparison we conducted quantitative assessments including: coefficients of variance, percentage differences between groups and standardized effect sizes calculated with Cohen's d . Across species and anatomical regions, the overall literature supports the presence of age‐related MN loss. Evidence is strongest in studies using robust counting approaches such as stereology, and appropriate MN markers. As age‐related MN degeneration is more heavily investigated and becomes a locus of intervention, future work must employ rigorous methodological standards to accurately determine the extent, regional specificity, and functional consequence of aging MN loss.

Effective neuromotor function may be considered the combination of activating motor units of a type, matching the desired force and fatigue requirements of a behaviour; and the timing of these activations in concert with motor units of other motor pools and/or with contextual/sensory cues. Although the former is well-established as being impaired with age (i.e., sarcopenia), the latter timing issues remain poorly defined in terms of aging. Indeed, sarcopenia may be considered the apotheosis of a degenerative process that first affects the timing and effective activation of motor units prior to overt weakness. We hypothesized that mild motor impairment (i.e., timing deficiencies and motor neuron synaptic pathology) would precede overt striated muscle weakness and motor neuron death. To test this hypothesis, we evaluated the neuromotor system for swallow, where striated muscle activities do not create torque across a joint (a confounder in aging studies) but pressure in the oropharynx and coordination with ventilation. We used female and male Fischer 344 rats (F344) at 6- (young), 18- (late middle-age) and 24-months-old (old) rats. Behavioral evaluations consisted of pharyngeal and thoracic pressures during naturalistic (0.5 ml oral water bolus) swallow. Synapse evaluations consisted of golgi-cox evaluation of nucleus ambiguus and hypoglossal motor neurons, and serial blockface electron microscopy evaluation of hypoglossal motor neurons. We assessed longitudinal (intrinsic) and genioglossus (extrinsic) muscle function and histopathology using platinum plate electrodes and immunohistochemistry for myosin heavy chain (MyHC) expression, respectively. Overall, we show that swallow timing abnormalities occur at late middle-age, contemporaneous with dendritic synaptic and mitochondrial deficits. Notably, motor neuron survival and muscle function and cross-sectional areas were unchanged in late middle- age, with sarcopenia and nucleus ambiguus and hypoglossal motor neuron death only apparent from old age. These results are consistent with the concept of “mild motor impairment” being a harbinger of striated muscle sarcopenia and frailty. There is every reason to suggest that these early motor timing deficits occur in spinal motor neurons. These results illustrate the importance motor neuron health has on maintenance of motor function into old age. Funding: This study was supported by R56 HL166204 & R01 AG086136 (MJF) This abstract was presented at the American Physiology Summit 2026 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.

Sarcopenia is the age-associated muscle weakness and atrophy. As this muscle weakens, so does the capacity for an animal to perform its associated behaviors. The tibialis anterior (TA) muscle is a lower leg muscle that is important for dorsiflexion during actions such as jumping and running. In humans and Fischer 344 rats, behavioral and pathophysiological assessments show spinal cord motor unit dysfunctions with age are primarily of fast fatigueable motor units, comprising larger MN and the type IIx/b fibers they innervate. In addition, mitochondrial dysfunctions are implicated in aging neuromotor dysfunction and muscle weakness, however, little fiber type specific knowledge exists regarding the function of aged mitochondria in type-identified muscle fibers. Based on the specific atrophy and reduced oxidative capacity (succinate dehydrogenase [SDH] enzymatic function) of type IIx/b fibers in respiratory muscle, we expected to observe type-specific atrophy and reduced SDH activity in type IIx/b TA fibers. In young (6-months-old, n=12) and old (24-months-old, n=12) female and male F344 rats, cut serial TA sections (8 µm) on a cryostat to evaluate type specific morphology and SDH. To determine CSA TA fibers were labelled with MyHCSlow (Novus NBP2 50299), MyHC2A (Novus N1551) and anti-Laminin antibodies, with alexa488, alexa555 and alexa408 secondaries, respectively. SDH was determined using 10 min incubation n 80 mM succinate with 1.5mM nitro blue tetrazolium, with precipitate imaged using a Nikon AR1 inverted microscope with a 579nm high pass filter. We observed a fiber type-specific atrophy of IIx/b fibers in old TA fibers. We also observed a marked reduction in TA SDH activity in old age, confined to type IIx/b fibers. These findings illustrate consistent fast fatigue resistant motor unit degeneration in old age. Strategies specifically targeted to these motor units are imperative in interventions of sarcopenia. Funding: This study was supported by R56 HL166204 (MJF), R01 AG086136 (MJF) and AG44615 (GCS) from the NIH. This abstract was presented at the American Physiology Summit 2026 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.

Brain derived neurotrophic factor (BDNF) signals through its high-affinity full-length tropomyosin receptor kinase-B (TrkB.FL) and plays role in maintaining the survival and function of phrenic motor neurons (PhMNs), neuromuscular transmission and synaptic activity. Disruption of this pathway is a hallmark of several neurodegenerative diseases associated with dysfunction and death of PhMNs with impaired mitochondrial function. BDNF/TrkB.FL signaling phosphorylates CREB at serine 133 (pCREBS133) and promotes its nuclear localization in cervical ventral horn of rats, however its downstream molecular pathway has not yet been established. Recently, we demonstrated that in human airway smooth muscle cells, pCREBS133, as a transcription factor, binds to the CRE sequence in the PGC1a promoter and activates its expression. PGC1a co-transcriptionally activates mitochondrial transcription factor A (TFAM) expression promoting mitochondrial DNA (mtDNA) replication and mitochondrial biogenesis. In the present study, we hypothesized that BDNF/TrkB.FL signaling promotes pCREBS133 phosphorylation and transcriptional activation of PGC1a and downstream expression of TFAM, leading to mitochondrial biogenesis. In this study, we employed a loss of function approach using a novel chemogenetic rat model (TrkBF616A) with a knock-in allele sensitive to the selective kinase inhibitor 1NMPP1 to inhibit TrkB.FL kinase activity. One group of female and male rats were treated intrathecally at C4 with either artificial cerebrospinal fluid (aCSF) or BDNF (450 ng/ 300 g b.w.; diluted in aCSF) for 40 mins, followed by collection of C3-C6 cervical ventral horn. In a second group, female and male rats were treated with 1NMPP1 (25 uM; i.p.) or the vehicle DMSO for 2 h. After 2 h, both groups were treated intrathecally with BDNF for 40 min and the C3-C6 cervical ventral horn collected. Intrathecal BDNF increased the pTrkB.FLY817 autophosphorylation and downstream phosphorylation of pCREBS133. Bioinformatic analysis identified a putative binding site of pCREBS133 on PGC1a promoter, which was confirmed by Chromatin immunoprecipitation (ChIP) assay. Consistent with this, BDNF treatment increased the expression of PGC1a and downstream TFAM, with an increase in mtDNA copy number. 1NMPP1 inhibition of TrkB.FL kinase (blunted BDNF-induced pTrkB.FLY817 phosphorylation) inhibited downstream activation of the pCREBS133/PGC1a/TFAM signaling cascade. Additionally, 1NMPP1 treatment blunted the BDNF-induced increase in mtDNA copy number. We conclude that BDNF/TrKB.FL signaling mediates mitochondrial biogenesis in the cervical ventral horn suggesting that interventions targeting this pathway could be a therapeutic strategy for neurodegenerative diseases. This research was funded by NIH Grants HL146114 & AG44615. This abstract was presented at the American Physiology Summit 2026 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 aerodigestive system (ventilation, swallow, speech and airway defense manoeuvres), age-related neuromotor declines contribute to the increased risk of aspiration pneumonia, a leading cause of mortality in the elderly. In the Fischer 344 (F344) rat model, we and others have identified that brainstem hypoglossal motor neuron (MN) degenerations and death contribute to motor dyscoordination and tongue muscle weakness, respectively. In F344 rats, both MNs and striated muscle exhibit morphological and functional deficits of mitochondria in aging, consistent with elderly human post mortem results. However, to date the dendritic and axonal compartments have been relatively ignored. In young (6-months old; n=6 rats) and old (24-months-old; n=6 rats) female and male F344 hypoglossal motor axons were assessed using serial block face scanning electron microscopy (SDFEM). Within myelinated hypoglossal motor axons and hypoglossal MN dendrites, mitochondrial volume density and morphology (mitochondrial CSA, form factor and aspect ratios) were assessed. In old age, mitochondrial volume density was reduced in dendrites and, surprisingly, increased in axons. Mitochondrial fragmentation (reduced CSA, form factors and aspect ratios was apparent in dendrites. In axons, old age mitochondria were larger, but more simplistic. We interpret these dendritic findings to be consistent with altered neural network function in old age. In the axons, excessive mitochondria may physically impair the anterograde and retrograde transit along the axons. Our results suggest that a more nuanced compartment-specific evaluation of mitochondrial structure and function is required to fully elucidate the pathophysiology underlying age-related neuromotor dysfunction. Funding for this research was provided by National Institutes of Health (NIH) grant AG086136 (MJF). This abstract was presented at the American Physiology Summit 2026 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.