Spinal motor neurons serve as the link between the nervous system and muscles. As the final common pathway of the neuromuscular system, they receive inputs from both higher-level controllers and afferent pathways. It is often assumed that spinal motor neurons are primarily driven by continuous common inputs (cCI) within different frequency bands. Within this framework, the motor neuron pool behaves as a linear amplifier of the cCI. This implies that the frequency content of descending and spinal oscillatory signals is preserved and faithfully transmitted to the muscles; thus, the spectral content at the output of the motor neuron pool corresponds to that of the cCI. However, this framework overlooks the possibility that motor neurons could also be driven by impulsive common inputs (iCI), which can induce synchronization among them and disrupt the linear transmission of other synaptic inputs at the pool level. To test this hypothesis, computational simulations and experimental data from two different human muscles were used to characterize different aspects related to motor neuron spiking synchronization at the pool level. Our findings suggest that, indeed, iCI can account for relevant features observed in experimental data such as the presence of synchronization events at the pool level. We also observed that such impulsive inputs can affect the linearity in the transmission of cCI by the motor neuron pool. This study represents pioneering indirect evidence of the existence of iCI as inputs to motor neurons. KEY POINTS: The current understanding of the motor control of voluntary movements assumes a continuous control, driven by oscillatory common signals. Some aspects of motor unit pool behaviour (particularly in terms of spiking synchronization and spectral content) typically observed in experimental recordings cannot be reproduced in simulations that only use continuous common inputs (cCI) to motor neurons. This study provides evidence indicating that spinal motor neurons receive a portion of their synaptic input in the form of impulsive common inputs (iCI) that synchronize their activity. The study also shows how such iCI can affect the linear transmission of other cCI by the motor neuron pool. These findings constitute a fundamental paradigm shift in the understanding of motor control and impact the development of interfaces that extract information from the activity of spinal motor neurons.
Neural circuits exhibit remarkable plasticity in response to varying intensities of sensory input. The temporal dynamics and cellular mechanisms underlying this plasticity are highly heterogeneous and possibly specific to individual circuits. Excessive noise exposure causes damage of peripheral auditory structures, such as cochlear hair cells and auditory nerve fibres, reducing afferent projection to downstream structures and initiating cascades of long-lasting compensatory changes in central auditory circuits. Amongst these changes, increased neuronal excitability, elevated spontaneous firing and increased neural gain were reported across multiple structures between the cochlear nucleus and auditory cortex. However, these findings primarily involved neurons that were responsive to sound onset (ON) and driven by excitation. Much less is known about the impact of noise exposure on neurons that are selectively activated by sound offset (OFF) and are driven by inhibition. We addressed this gap in knowledge by investigating the effects of noise exposure on intrinsic membrane properties, synaptic input patterns and sound-evoked activity in superior paraolivary nucleus (SPN) neurons, which are a population of neurons specialized for encoding sound offset. Immediately after noise exposure, SPN neurons were unresponsive to sound offset. Within the next 24 h, we observed a significant increase in the number of inhibitory synaptic terminals impinging upon SPN neurons, which was corroborated by elevated frequencies and amplitudes of inhibitory postsynaptic currents. At the same time, SPN neurons exhibited higher intrinsic excitability. Together, these changes encouraged recovery of sound-evoked OFF responses 24 h following noise exposure, suggesting circuit-specific compensatory mechanisms that enable sound OFF encoding soon after peripheral auditory insult. KEY POINTS: Sound-offset (OFF) responses mark the critical temporal boundary when a sound terminates; this enables encoding of sound duration and the detection of gaps in sounds and calls. In the mouse model, OFF responses are generated de novo in the superior paraolivary nucleus by combining sound-evoked inhibitory input with the intrinsic membrane properties of the neurons. The impact of noise over-exposure on these OFF responses and its implications for subsequent auditory processing is not well understood. Combining patch-clamp recording, immunohistochemistry and in vivo electrophysiology, we show that superior paraolivary nucleus neurons exhibit increased excitability and enhanced inhibition following noise over-exposure. These compensatory changes help to mediate early recovery of sound OFF responses to loud stimuli, despite the loss of auditory sensitivity at lower sound intensities.
Renal juxtaglomerular renin-producing cells and preglomerular vascular smooth muscle cells (VSMCs) are specialized pericytes with notable plasticity. Preglomerular VSMCs can convert to renin-producing cells during severe hypotension or salt depletion, and renin cells can transform into erythropoietin (EPO)-producing cells when hypoxia‑inducible factor (HIF)-2α is stabilized through deletion of prolyl-4-hydroxylases (PHD) 2 and 3. These findings raise the question of whether PHD2 and PHD3 likewise regulate the endocrine plasticity of preglomerular VSMCs. To investigate the role of PHD2 and/or PHD3 in (preglomerular) contractile pericytes, inducible mouse models with smooth muscle myosin heavy chain (SMMHC)-specific deletion of PHD2 and/or PHD3 were examined under basal conditions or after stimulation of renin production by treating the mice with a low-salt diet and angiotensin converting enzyme inhibitor enalapril (LSE). At baseline, none of the deletions altered renin production or induced EPO expression in preglomerular pericyte-like VSMCs, despite HIF-2α stabilization in PHD2/PHD3-deficient mice. However, HIF-2α stabilization resulting from PHD2 or PHD2/PHD3 deletion triggered EPO production in interstitial SMMHC+ contractile pericytes. LSE treatment induced renin in VSMCs and extraglomerular mesangial cells of control, SMMHCCreERT2 PHD2ff and SMMHCCreERT2 PHD3ff mice. In contrast, VSMCs of PHD2/PHD3-deficient mice produced EPO rather than renin, while renin induction persisted only in mesangial cells. Notably, this LSE-induced EPO production was reversible despite ongoing HIF-2α stabilization. Transcriptional changes indicated a shift in PHD2/PHD3-deficient VSMCs from a contractile/renin cell-like to a contractile/EPO cell-like signature. These findings indicate that HIF-2α stabilization determines the endocrine product of preglomerular VSMCs and interstitial pericytes. Notably, loss of PHD2/PHD3 does not compromise the plasticity of VSMCs to reversibly adopt endocrine functions. KEY POINTS: Smooth muscle myosin heavy chain (SMMHC)-specific deletion of the prolyl‑4‑hydroxylases PHD2 and PHD3 stabilized hypoxia‑inducible factor (HIF)‑2α in preglomerular pericyte‑like vascular smooth muscle cells (VSMCs), prompting a transcriptional shift from a contractile/renin cell‑like toward a more contractile/EPO cell‑like signature without activating erythropoietin (EPO) transcription. A reduction in systolic blood pressure through treatment with low-salt diet and angiotensin converting enzyme inhibitor enalapril induced EPO synthesis instead of renin in preglomerular PHD2/PHD3-deficient VSMCs. Transformation of preglomerular VSMCs into EPO-producing cells was reversible despite persistent HIF-2α stabilization. SMMHC cell-specific deletion of PHD2 and PHD2/PHD3 activated EPO production in interstitial contractile pericytes independent of systolic blood pressure. Short‑term HIF‑2α stabilization was insufficient to induce EPO production in preglomerular VSMCs or contractile pericytes. Τaken together these findings demonstrate that HIF-2α stabilization governs the endocrine output of preglomerular VSMCs and interstitial pericytes. Notably, the loss of PHD2/PHD3 does not impair the capacity of VSMCs to reversibly assume endocrine functions.
The voltage-gated Nav1.7 and Nav1.8 channels are essential to transmit acute and chronic pain. Increased trafficking of Nav1.7 and Na1.8 channels to the membrane of sensory neurons is a critical mechanism of pain sensitization. However the mechanisms responsible for the trafficking of Nav1.7 and Nav1.8 channels remain unclear. We found that acute nociception and heat and mechanical hyperalgesia induced by the activation of nociceptive TRPV1 and TRPA1 channels were markedly reduced in IQGAP1-deficient mice. The basal excitability of sensory neurons was also significantly reduced in the absence of IQGAP1. Correspondingly the deletion of IQGAP1 reduced the basal membrane expression of Nav1.7 and moreover prevented enhanced trafficking and sensitization of Nav1.7 and Nav1.8 channels in sensory neurons induced by inflammatory mediators (IM). Heat hyperalgesia, mechanical and cold allodynia in nerve injury induced neuropathic pain mediated by Nav1.7 and Nav1.8 channels, respectively, were also prevented in IQGAP1-deficient mice. IQGAP1 thus governs the trafficking and potentiation of both Nav1.7 and Nav1.8 channels and could be exploited for therapeutic interventions for the treatment of acute and chronic pain. KEY POINTS: The enhanced activities of voltage-gated Na+ channels Nav1.7 and Nav1.8 in sensory neurons underpin acute and chronic pain. Enhanced activities of Nav1.7 and Nav1.8 channels are due to an increased number of these channels inserted into the plasma membrane of sensory neurons. This study reveals that the scaffold protein IQ motif containing GTPase activating protein 1 (IQGAP1) mediates forward membrane trafficking of Nav1.7/Nav1.8 channels and resultant functional enhancement of these channels. Enhanced pain caused by inflammatory mediators and nerve injury is reduced in the absence of IQGAP1. Our findings elucidate the trafficking mechanisms of Nav1.7 and Nav1.8 channels and suggest IQGPA1 as an alternative treatment option for acute and chronic pain.
How neuroinflammation is altered at high altitude (HA) is equivocal, and it is unclear whether and how exercise alters this inflammatory response. We tested the hypothesis that maximal exercise at HA exacerbates the neuroinflammatory response. Healthy adults (n = 12, 6/6 females/males) completed 60 min of steady-state semi-recumbent low-intensity cycling exercise and an incremental maximal test at sea level (SL) and following 6-8 days at 3800 m a.s.l. (HA). Radial arterial and internal jugular venous (IJV) sampling and duplex volumetric blood flow of the internal carotid and vertebral arteries was used to assess trans-cerebral exchange of cytokines at rest and 5 min after maximal exercise (POST-MAX-5). Arterial-IJV uptake/release of cytokines at rest was not different between altitudes. Arterial leucocytes were increased by 57%-66% at POST-MAX-5 at both SL (P < 0.001) and HA (P < 0.001); exercise × altitude, P = 0.433, with no difference between arterial and IJV. Apart from a trivial absolute increase in arterial interleukin-6 at HA (Δ +0.62 pg ml-1, 95% confidence interval [0.10, 1.15], P = 0.022), the remaining 16 of the 17 measured systemic cytokines were unaffected by maximal exercise at both altitudes (P > 0.05). There was a shift towards trans-cerebral net release by -1.3 to -8.2 pg ml-1 (P < 0.05) of interleukin-6, interleukin-8, monocyte chemoattractant protein-1 and macrophage inflammatory protein-1 beta at POST-MAX-5 at both altitudes. The release of these cytokines occurred independently of systemic inflammation and altitude exposure. The results of this study indicate that partial acclimatization at 3800 m a.s.l. does not alter the physiological release of cytokines from the brain in response to maximal exercise. KEY POINTS: Systemic inflammation is altered at high altitude, but it is unclear how exercise at altitude affects inflammation in the brain. We tested the hypothesis that maximal exercise will augment the cerebrovascular inflammatory response at high altitude. Exposure to 6-8 days at high altitude did not evoke systemic or brain inflammation at rest and did not exacerbate responses to maximal exercise. There was a shift towards net release of interleukin-6, interleukin-8, monocyte chemoattractant protein-1 and macrophage inflammatory protein-1 beta following maximal exercise at both altitudes, driven by an increase in internal jugular venous concentration. These new results show that the adaptive physiological response to exercise involves net release of select cytokines from the brain; this occurred in the absence of systemic inflammation and irrespective of high altitude.
Cardiotoxicity can be a result of the action of various therapeutic drugs, including anti-cancer drugs, and can pose a greater risk to patients' health than the underlying disease being treated. As cardiac function relies on the interplay between different cell types, dysfunction in cell-cell crosstalk can promote disease-associated mechanisms. Extracellular vesicles (EVs) are lipid bilayer structures that provide a means of cell-cell communication via delivery of bioactive cargo. EVs contribute to both physiology and pathophysiology, having recently been implicated in the pathogenesis of cardiovascular diseases (CVDs). Cardiotoxicity displays pathological similarities to those seen in numerous CVDs, suggesting a potential role for EVs in this process. This role has been confirmed in studies on the anti-cancer drug doxorubicin, observing that EVs generated by doxorubicin-treated cancer cells can drive cardiotoxicity in healthy cells present in the cardiovascular system. Conversely, EVs have also shown therapeutic potential in treating cardiovascular pathologies. EVs, such as those originating from stem cells, have been observed to disrupt pathogenesis of CVD and doxorubicin-induced cardiotoxicity, suggesting potential for EV-based therapeutics. As current therapeutic strategies tackling cardiotoxicity treat late-stage pathological changes and have limited long-term efficacy, therapeutics targeting mechanisms that initiate and drive cardiotoxicity may be more effective in preserving cardiac function. Overall, this review demonstrates the potential contribution of EVs in cardiovascular (patho)physiology through mediating cell-cell communication. This review aims to highlight the need for further study to fully understand the significance of EVs in driving drug-induced cardiotoxicity and includes suggestions for methods of establishing accessible EV-based therapeutics for at-risk patients.
The characterization of skeletal muscle phenotypes in diving populations remains one of the least explored domains of breath-hold physiology, representing a critical gap in our understanding of how skeletal muscle adapts to the unique demands of breath-hold diving. Accordingly the present study investigated specific markers of skeletal muscle structure and metabolism in competitive breath-hold divers. Twenty males volunteered to participate in this study (10 competitive breath-hold divers; 10 non-divers), matched for age, body size and whole-body aerobic capacity ( V ̇ O 2 max ${\dot{\mathrm{V}}}\rm{O}_{\rm{2max}}$ ). A percutaneous skeletal muscle biopsy was obtained from the m. vastus lateralis to quantify capillarization, fibre-type distribution (i.e. types I, IIa and II other), protein content of mitochondrial complexes, monocarboxylate transporter (MCT) isoforms and citrate synthase activity. MCT4 content was 28% higher in breath-hold divers compared to non-divers (P = 0.020), whereas MCT1 and citrate synthase activity showed no between-group differences (P ≥ 0.161). Complex V content was higher in the non-divers (P = 0.049), whereas no between-group differences were noted for complexes I, II, III and IV (P ≥ 0.253). Capillarization was significantly higher in breath-hold divers (P ≤ 0.048), whereas fibre-type distribution did not differ between groups (P = 0.999). Competitive breath-hold divers exhibited skeletal muscle characteristics indicative of enhanced blood-muscle exchange capacity and augmented lactate efflux potential. Such adaptations may confer an advantage during prolonged breath-holds by preserving glycolytic function and maintaining redox homeostasis. In recovery these traits likely facilitate more efficient clearance of metabolic byproducts. KEY POINTS: The skeletal muscle phenotype of breath-hold divers remains poorly characterized, limiting understanding of how skeletal muscle adapts to the physiological demands of prolonged breath-holding. This study compared skeletal muscle structure and metabolic markers between competitive breath-hold divers and non-diving controls matched for age, body size and whole-body aerobic capacity. Breath-hold divers exhibited greater skeletal muscle capillarization, indicating an enhanced capacity for blood-muscle exchange. They also showed an increased potential to remove lactate from skeletal muscle tissue. These adaptations likely support sustained glycolytic function and redox balance during prolonged breath-holds, while facilitating more efficient clearance of metabolic byproducts during recovery.
Propionate, a gut microbiota-derived short-chain fatty acid, influences fetal development and postnatal metabolic programming. Although the fetus lacks microbiota and endogenous propionate production, human pregnancies show a fetal-to-maternal propionate concentration ratio greater than unity, suggesting concentrative transport across the placenta. However, its underlying mechanism remains undefined. The present study aimed to identify the transporter responsible for transplacental transport of propionate across the syncytiotrophoblast (SynT) layer. Transporter knockdown in human choriocarcinoma JEG-3 cells revealed that MCT1 (SLC16A1) silencing reduced [3H]propionate uptake, whereas knockdown of other anion transporters had no effect. Functional assays using Xenopus oocytes demonstrated that the expression of human MCT1, but not MCT4 (SLC16A3), increased [3H]propionate transport. In human trophoblast stem cell (hTSC)-derived SynT, [3H]propionate uptake was pH-dependent and significantly inhibited by MCT1-selective inhibitors. Subsequently, to evaluate transcellular transport, we performed quantitative permeability assays using a hTSC-derived placental barrier model. [3H]Propionate permeability was significantly higher than that of [14C]d-mannitol, a paracellular marker. MCT1 inhibition reduced [3H]propionate permeability in both apical-to-basal and opposite directions, whereas MCT4 inhibition had minimal effects. Notably, the hTSC-derived model exhibited a directional bias in [3H]propionate transfer, reflecting the fetal-directed enrichment observed in vivo. Mathematical model analysis further indicated that MCT1 functions at both the apical and basal membranes to facilitate bidirectional transport of propionate. Together, these findings identify MCT1 as the predominant mediator of propionate transfer across the human SynT layer, providing mechanistic insights into how the placenta governs fetal exposure to maternal microbiota-derived metabolites. KEY POINTS: The fetus relies on maternal-derived propionate for development, but the molecular mechanism responsible for its concentrative transport across the human placenta remains undefined. Using multiple human trophoblast models and functional expression assays, we identified MCT1, but not MCT4, as the primary mediator of propionate transport. A human trophoblast stem cell-derived placental barrier model successfully exhibited a directional bias in propionate transfer, reflecting the fetal-directed enrichment observed in vivo. Mathematical modelling of the permeability data from the human trophoblast stem cell-derived barrier model indicates that MCT1 functions at both the apical and basal membranes of the syncytiotrophoblast to facilitate bidirectional transport. These findings establish MCT1 as a key gateway for transplacental propionate transfer, providing mechanistic insights into how the placenta regulates fetal exposure to maternal microbiota-derived metabolites.
Slow skeletal muscles maintain posture and produce graded movement at low metabolic cost. ATP utilization during fixed-end contractions is typically five times slower in slow muscles than in fast muscles from the same species. Mechanical measurements previously suggested that more myosin motors are attached to thin filaments during contraction of slow muscle, which seems incompatible with its high efficiency. We therefore used small-angle X-ray diffraction to provide a structural estimate of the fraction of myosin motors attached to thin filaments in slow muscle. The X-ray signals associated with myosin binding to actin indicate that only ∼10% of myosin motors are actin bound during fixed-end tetani of rat soleus slow muscles, compared with ∼25% in mouse extensor digitorum longus fast muscle. Moreover, X-ray signals associated with the helical organization of OFF myosin motors in the thick filaments show that ∼70% of myosin motors remain in the OFF conformation during tetanic contraction of rat soleus muscle, compared with only 30% in mouse extensor digitorum longus muscle. The much slower force development in soleus muscle also allowed clear separation of early structural changes in thick filaments on activation, some of which are distinct from those reported previously in fast muscles. Moreover, the early structural changes in soleus muscle have about the same amplitude in a twitch and a tetanus, suggesting that they are triggered by thin filament activation rather than thick filament stress and implying a fast signalling pathway between thin and thick filaments. KEY POINTS: The interaction between myosin motors and actin filaments in slow skeletal muscles maintains posture and produces graded movement at low metabolic cost. Mechanical studies have suggested that more myosin motors are attached to actin filaments during isometric contraction of slow than fast muscle, but this seems incompatible with its high efficiency. We used X-ray diffraction to show that there are fewer myosin motors attached to actin in slow muscle than in fast muscle because more motors are sequestered on the myosin filament. The slower force development in slow muscle also allowed us to isolate and characterize fast changes in myosin motor conformation associated with activation of the actin filaments.
A functional blood-brain barrier (BBB) is essential for CNS homeostasis, and its disruption is an early feature of both acute brain injury and chronic neurodegenerative disorders. Hypoxia induces BBB breakdown by triggering endothelial dysfunction, oxidative stress, metabolic dysregulation and thrombo-inflammatory signalling that compromise barrier integrity. However, strategies that restore BBB function remain limited. Here, we investigated whether photobiomodulation (PBM), a non-invasive light therapy, can rescue BBB dysfunction following acute hypoxic stress. Using a multicellular in vitro BBB model comprising immortalised human brain microvascular endothelial cells, pericytes and astrocytes, we induced hypoxic injury (6 h, 1% O2) and applied three PBM treatments during recovery. Hypoxia significantly reduced transendothelial electrical resistance (TEER), whereas PBM restored barrier function in endothelial monocultures and tri-cultures. Endothelial cells exhibited the most pronounced hypoxic response, characterised by increased expression of hypoxia-inducible factor-1α (HIF-1α), plasminogen activator inhibitor-1 and von Willebrand factor (vWF), all attenuated by PBM. Importantly, small interfering RNA-mediated knockdown of vWF partially recapitulated PBM-induced restoration of barrier integrity, identifying endothelial vWF as a mediator of recovery. PBM also reduced reactive oxygen species in hypoxic astrocytes and pericytes, indicating co-ordinated multicellular modulation. Together, these findings demonstrate that PBM restores BBB integrity following hypoxic insult by modulating endothelial thrombo-inflammatory signalling at the same time as reducing oxidative stress in glial cells. Rather than acting as a non-specific cytoprotective stimulus, PBM engages molecular pathways linked to endothelial activation. This work establishes a mechanistically informed platform for investigating BBB repair and highlights PBM as a strategy to protect vascular integrity in hypoxia-associated neurological disorders. KEY POINTS: Hypoxia is a major driver of blood-brain barrier (BBB) dysfunction, yet there are currently no targeted therapies that directly restore barrier integrity. Photobiomodulation (PBM) is a non-invasive low-level light intervention known to facilitate mitochondrial function and cellular stress responses. In a human in vitro BBB model, repeated PBM treatment restored transendothelial electrical resistance (TEER) 24 and 48 h after hypoxic injury, with endothelial rescue linked to downregulation of von Willebrand factor (vWF). PBM modulated oxidative stress, hypoxia signalling and thrombo-inflammatory pathways across endothelial cells, astrocytes and pericytes. These findings support PBM-driven modulation of endothelial signalling as a potential strategy to restore BBB integrity in hypoxia-associated neurological conditions.
Gastric muscles were obtained from obese patients with no other underlying morbidities undergoing vertical sleeve gastrectomy (VSG). Quantitative electrophysiological techniques were used to characterize the ionic mechanisms underlying electrical slow waves in muscles of the gastric antrum. Thin muscular sheets were prepared using vibratome sectioning to characterize the electrical activity through the thickness of the tunica muscularis. Two distinct pacemaker regions were identified: large-amplitude, long-duration slow waves occurred at low frequency in longitudinal muscle (LM) near the serosa; higher-frequency, shorter-duration slow waves were recorded in muscle near the myenteric plexus and throughout the circular muscle (CM). The higher-frequency pacemaker dominated activity and generated phasic contractions in intact muscles. The upstroke depolarization of slow waves depended predominantly on T-type Ca2+ channels, but CaV1.2 and CaV1.3 L-type channels also participated. Ca2+ entry during the upstroke appeared to initiate Ca2+-induced Ca2+ release that was sustained for several seconds, activating Ca2+-activated Cl- channels (ANO1). Ca2+ release occurred from stores loaded by sarco/endoplasmic reticulum Ca2+-ATPase (SERCA), and both IP3 and ryanodine receptors were involved in the Ca2+ release that activated ANO1. The elevation of intracellular Ca2+ required to maintain the activation of ANO1 channels through the plateau phase relied upon sustained Ca2+ entry through L-type Ca2+ channels and Na+/Ca2+ exchange operating in reverse mode. Stores of Ca2+ were maintained over time by store-operated Ca2+ entry mediated by ORAI. Slow waves generated the phasic contractions underlying gastric peristalsis. Thus this study provides mechanistic information about the electrophysiology underlying gastric motility. KEY POINTS: What is known about gastric electrophysiology is primarily deduced from animal studies and extracellular recordings from human patients. It is difficult to determine the ionic mechanisms underlying components of pacemaker activity from extracellular recordings. Using quantitative intracellular microelectrode recordings two distinct pacemaker regions were identified in the human gastric antrum: large, long-duration, low-frequency slow waves in the longitudinal muscle and higher-frequency, shorter-duration slow waves near the myenteric plexus and throughout the circular muscle. The higher-frequency pacemaker dominated activity and generated phasic contractions in intact muscles. Pacemaker activity involved a complex series of events, including (i) Ca2+ entry through voltage-dependent Ca2+ channels, (ii) Ca2+ release from intracellular sarco/endoplasmic reticulum Ca2+-ATPase stores, (iii) activation of ANO1 and (iv) sustained increase in intracellular Ca2+ via a Na+/Ca2+ exchanger operating in reverse mode.
The medial prefrontal cortex (mPFC) undergoes extensive GABAergic interneuron maturation during adolescence, a process that establishes excitatory-inhibitory balance and supports adult cognitive function. Disruptions during this critical developmental period contribute to neuropsychiatric disorders, yet the enduring consequences for adult synaptic plasticity remain poorly understood. Here we examine how adolescent NMDA receptor (NMDAR) hypofunction impacts adult mPFC transmission and plasticity. We exposed adolescent mice to subanaesthetic ketamine and performed whole-cell patch-clamp recordings in adulthood to examine excitatory and inhibitory synaptic currents and spike timing-dependent plasticity (STDP). Ketamine-exposed mice exhibited persistent GABAergic transmission deficits in pyramidal neurons (PyNs), as evidenced by reduced spontaneous and miniature IPSC frequencies and elevated paired-pulse ratios, consistent with impaired presynaptic GABA release and reduced functional output of parvalbumin-positive interneurons (PV-INs). PyN STDP was altered in ketamine-exposed mice, with spike pairings in the post-before-pre order inducing potentiation, in contrast to the synaptic depression observed in vehicle mice. Together, these results demonstrate that adolescent NMDAR hypofunction produces enduring impairments in PV-IN-mediated inhibitory transmission and disrupts the bidirectional expression of STDP in the adult mPFC. This shift in plasticity rules reflects a loss of inhibitory control over synaptic integration and indicates that developmental NMDAR disruption produces persistent alterations in cortical circuit function. Together, these findings provide mechanistic insight into how adolescent NMDAR hypofunction leads to enduring circuit dysfunction, with relevance to neurodevelopmental disorders emerging during adolescence. KEY POINTS: Adolescence is a critical period for GABAergic maturation in the medial prefrontal cortex (mPFC). Adolescent ketamine exposure reverses adult mPFC spike timing-dependent plasticity from depression to potentiation. Adolescent ketamine exposure impairs GABAergic transmission in adulthood. Adolescent ketamine exposure reduces the functional output of parvalbumin-positive interneurons. These findings demonstrate that adolescent NMDA receptor hypofunction disrupts adult prefrontal inhibitory balance and reverses plasticity polarity.
High-altitude hypoxia constrains tissue O2 supply, but several high-altitude populations have evolved adaptations to overcome this challenge. Evolved increases in haemoglobin-O2 (Hb-O2) affinity are pervasive across high-altitude taxa, but the influence of such increases on aerobic capacity in hypoxia remains contentious. The influence of Hb-O2 affinity could depend on the capacity to extract O2 from the blood, but this possibility is poorly understood. We examined this issue in deer mice (Peromyscus maniculatus), which are found from sea level to >4300 m elevation in the Rocky Mountains. Mice from populations native to high and low altitudes were born and raised in captivity. Low-altitude mice were acclimated to warm (25°C) normoxia and high-altitude mice were acclimated to cold (5°C) hypoxia (∼12 kPa O2), creating two groups with distinct capacities for O2 transport. Aerobic capacity for thermogenesis was measured in hypoxia after each of three pharmacological treatments: saline (control), efaproxiral (decreases Hb-O2 affinity) and cyanate (increases Hb-O2 affinity). High-altitude mice had greater aerobic capacity in hypoxia, in association with higher arterial O2 saturation ( S a O 2 ${{S}_{{\mathrm{a}}{{{\mathrm{O}}}_2}}}$ ) and lower P50 (O2 pressure at 50% Hb saturation) in most conditions. The P50 at which aerobic capacity was greatest was lower in high-altitude mice than in low-altitude mice. High-altitude mice also had greater uncoupling protein 1 (UCP-1) content in brown adipose tissue and greater cytochrome oxidase activity in gastrocnemius muscle. These results suggest that optimal Hb-O2 affinity and S a O 2 ${{S}_{{\mathrm{a}}{{{\mathrm{O}}}_2}}}$ are greater in high-altitude mice, in association with a greater capacity to extract and consume O2 in thermogenic tissues. KEY POINTS: Evolved increases in haemoglobin-O2 affinity are pervasive across high-altitude taxa, but the influence of such increases on aerobic capacity in hypoxia remains contentious. We examined whether the influence of haemoglobin-O2 affinity on aerobic capacity for thermogenesis is altered in high-altitude deer mice. Using pharmacological treatments to manipulate haemoglobin-O2 affinity, we found that aerobic capacity in hypoxia was greatest at higher affinities in high-altitude mice than in low-altitude mice. Skeletal muscle and brown adipose tissue had more oxidative and thermogenic phenotypes in high-altitude mice. These results suggest that the optimal haemoglobin-O2 affinity in hypoxia is greater in high-altitude deer mice, potentially resulting from a greater capacity to extract and consume O2 in active tissues.
Gastrointestinal smooth muscle cell excitability is regulated by a syncytium of smooth muscle cells, interstitial cells of Cajal (ICCs) and platelet-derived growth factor receptor α+ cells. The mechanism of Ca2+ upregulation, such as Ca2+-activated Cl- channel (CaCC) for characteristic pacemaker activity of ICCs, is supported by the interaction between the endoplasmic reticulum as the Ca2+ source, the plasma membrane providing the membrane activity and mitochondria as a buffer. A concept of 'microdomains' consisting of these endoplasmic reticulum, plasma membrane and mitochondrial components has been suggested based on electrophysiological studies, laser microscopy and transmission electron microscopy. However, their entire structure and function cannot be understood without detailed 3-D information. In this article, 3-D analysis of the microdomain on CaCCs is reported to present a new interpretation of characteristic excitable Ca2+-dependent mechanisms. Novel volume-electron microscopy (EM) techniques, such as focused ion beam/scanning electron microscopy (FIB/SEM), are powerful tools to understand these mechanisms. These techniques can show cellular membrane contacts in sheet structures, and can calculate geometrical features, such as distance, surface area and volume. Geometric analysis of organelles and membranes with FIB/SEM represents a novel study of the gastrointestinal tract. Conventional transmission EM images and immunohistochemistry of new ICC subtypes are reviewed with respect to the relationship between anatomical and physiological functions. 3-D analysis of endoplasmic reticulum in motor neurons is also summarized as an example of other excitable cell types analysed by FIB/SEM.
Psychological stress is a recognised, yet mechanistically unresolved, risk factor for cardiovascular disease (CVD) partly through its association with a hypercoagulable state. Free radical-mediated oxidative stress has been proposed as a key upstream driver of this haemostatic imbalance. In this randomised cross-over study we investigated whether acute psychological stress promotes systemic radical formation and prothrombotic alterations in clot microstructure in eight healthy males. The Trier Social Stress Test was used to induce psychological stress. Antecubital venous blood was collected to measure the ascorbate free radical (A•-, electron paramagnetic resonance spectroscopy) and clot microstructure (Df, Fourier transform rheology), alongside standard coagulometry. Compared with the control condition (quiet sitting), psychological stress increased A•- (P = 0.042) and Df (P = 0.008), the latter reflecting larger, denser and more fibrin-rich networks. We also observed selective shortening of activated partial thromboplastin time (aPTT) (P = 0.018), indicating activation of the intrinsic coagulation pathway. This study provides the first in vivo evidence that acute psychological stress triggers systemic free radical formation and drives prothrombotic remodelling of clot architecture. These findings identify oxidative stress as a mechanistic link between psychological stress and CVD risk and highlight it as a compelling target for prevention and therapy. KEY POINTS: Although psychological stress is a recognised risk factor for cardiovascular disease (CVD), its mechanistic association with a hypercoagulable state remains unresolved. Acute psychological stress, induced by the Trier Social Stress Test (TSST), significantly increased systemic free radical formation, as measured by elevated levels of ascorbate free radical (A•-) via electron paramagnetic resonance (EPR) spectroscopy, and was associated with the formation of larger, denser, more fibrin-rich clots confirmed by increased fractal dimension (Df). The concurrent increase in A•- and Df suggests that free radical-mediated oxidative stress is an upstream driver of psychological stress-induced activation of haemostasis, specifically altering clot quality. The TSST selectively shortened activated partial thromboplastin time (aPTT), indicating activation of the intrinsic/contact coagulation pathway. There were no changes in prothrombin time (PT) or D-dimer, suggesting haemostatic activation occurred without engaging fibrinolysis. The findings demonstrate that even a brief episode of emotional stress can increase thrombotic potential in healthy individuals and identify oxidative stress as a key mechanistic link and a potential therapeutic target for treating stress-related CVD.
The melanocortin 4 receptor (MC4R) is a key component of the leptin-melanocortin system. Recent studies have started to expand our understanding of the role of MC4R beyond energy balance and sexual behaviour, exploring its potential influence on respiratory function, particularly in the context of obesity hypoventilation syndrome and obstructive sleep apnoea. The MC4R pathway is implicated in respiratory control. MC4R is expressed on the CO2-sensing neurons of the retrotrapezoid nucleus, and activation of the MC4R+ neurons in the retrotrapezoid nucleus augments the hypercapnic ventilatory response. MC4R is present at other sites of the neural respiratory network, where this receptor might also be involved in ventilatory control. The US Food and Drug Administration approved the MC4R agonist setmelanotide, which has been used effectively for the treatment of obesity involving the leptin-melanocortin pathway. Setmelanotide is also a promising treatment option for obesity hypoventilation syndrome and obstructive sleep apnoea, particularly in people with obesity caused by specific genetic disorders, including POMC, LEPRb, MC4R and other associated gene mutations. STATEMENT OF SIGNIFICANCE: Recent findings suggest that targeting the melanocortin 4 receptor (MC4R) might offer a new treatment approach for sleep-disordered breathing, particularly obesity hypoventilation syndrome (OHS). Previous literature suggests that genetic mutations in the MC4R pathway and pharmacological inhibitors of MC4R induce hypoventilation. New findings demonstrate that the MC4R agonist setmelanotide increases minute ventilation during sleep and wakefulness, reduces apnoeas during sleep and enhances the hypercapnic ventilatory response. These effects have been observed in animal models and might have implications for treating patients with OHS and obstructive sleep apnoea.
Understanding how pharmaceuticals cross the placenta is central to balancing effective maternal therapy with fetal safety. Computational and physiologically based pharmacokinetic models are powerful tools for predicting fetal drug exposure, but their accuracy depends on identifying the rate-limiting determinants of placental transfer. These include placental anatomy, the routes available for paracellular diffusion, and the cellular localisation and membrane polarisation of drug transporters. The placenta both limits and mediates fetal exposure: it clears pharmaceuticals from the fetal circulation yet provides the principal route by which drugs reach the fetus. Transfer depends on a drug's physicochemical properties, especially molecular size and lipid solubility, and the presence of specific transport mechanisms. Net fetal exposure reflects the combined effects of diffusion and transporter-mediated fluxes, plasma protein affinity, with a smaller contribution from placental metabolism. At the anatomical level recent advances in our understanding of placental ultrastructure have provided new routes allowing fetal exposure via diffusion. At the molecular level, defining transporter expression and polarisation is challenging and the literature is often inconsistent, reflecting methodological limitations. Single-cell and single-nucleus transcriptomics provide valuable insights into cell-specific gene expression, although not necessarily protein localisation. Spatial mass spectrometry offers complementary information on protein abundance and polarity but remains limited by resolution. Effective models require a strong foundation in physiology and anatomy. A key limitation to modelling placental drug transfer is clearly determining the cellular localisation and membrane polarisation of drug transporters, and addressing this question is key to developing more effective predictive models.
Chronic hypoxia during pregnancy increases uterine artery L-type CaV1.2 channel currents and uterine vascular resistance in pregnant ewes. In the present study, we tested the hypothesis that microRNA-210-mediated mitochondrial oxidative stress contributes to gestational hypoxia-induced CaV1.2 channel hyperactivity of uterine arteries. In a pregnant sheep model acclimatized to high-altitude hypoxia, we found that knockdown of endogenous microRNA-210 with microRNA-210 locked nucleic acid diminished hypoxia-induced CaV1.2 hyperactivity in uterine arteries. Accordingly, microRNA-210 mimic recapitulated the effect of hypoxia to increase CaV1.2 activity and window currents of uterine arteries from normoxic pregnant ewes. Mechanistically, we revealed that microRNA-210 mediated gestational hypoxia-induced suppression of mitochondrial respiration and increase of mitochondrial reactive oxygen species in uterine arteries. We provided evidence that MitoQ, a mitochondria-targeted antioxidant, blocked both gestational hypoxia- and microRNA-210-mediated CaV1.2 hyperactivity in uterine arteries from pregnant ewes. In addition, microRNA-210 significantly increased phenylephrine-induced vasoconstriction of uterine arteries of pregnant ewes, an effect that was inhibited by MitoQ. Thus, our study provides new evidence of a mechanistic link of microRNA-210-mediated mitochondrial oxidative stress in gestational hypoxia-induced L-type CaV1.2 channel hyperactivity of uterine arteries in an animal model of pregnant sheep and reveals potential targets for therapeutic interventions of pregnancy complications associated with chronic hypoxia. KEY POINTS: Knockdown of endogenous microRNA-210 with microRNA-210 locked nucleic acid diminished hypoxia-induced CaV1.2 hyperactivity in uterine arteries. MicroRNA-210 mimic recapitulated the effect of hypoxia to increase CaV1.2 activity and window currents of uterine arteries from normoxic pregnant ewes. MicroRNA-210 mediated gestational hypoxia-induced suppression of mitochondrial respiration and increase of mitochondrial reactive oxygen species in uterine arteries. MitoQ, a mitochondria-targeted antioxidant, blocked both gestational hypoxia- and microRNA-210-mediated CaV1.2 hyperactivity in uterine arteries from pregnant ewes. MicroRNA-210 significantly increased phenylephrine-induced vasoconstriction of uterine arteries of pregnant ewes, an effect that was inhibited by MitoQ.
The human heart beats 60-80 times a minute, which can amount to more than 3 billion heartbeats in one's lifetime. Each heartbeat is initiated by the sinoatrial node (SAN), a highly complex structure consisting of specialized cells that spontaneously fire action potentials (APs), propagating throughout the heart. Its automaticity is orchestrated by ion channels and transporters that contribute to the membrane and Ca2+ clocks, collectively known as the 'coupled clock'. Their activity is tightly regulated by autonomic and hormonal signalling pathways, most prominently β-adrenergic receptor (β-AR) signalling, which increases heart rate via activation of adenylyl cyclase (AC) and subsequent production of 3',5'-cyclic adenosine monophosphate (cAMP). In contrast, parasympathetic signalling through muscarinic M2 receptors reduces cAMP levels and activates inwardly rectifying K+ currents, thereby slowing pacemaker activity. The current topical review discusses recent literature encompassing the mechanisms of SAN regulation in health and disease, including cardiac arrhythmia syndrome such as catecholaminergic polymorphic ventricular arrhythmia, autoimmune cardiac ion channelopathies, and SAN dysfunction in heart failure (HF). SAN dysfunction in HF frequently manifests as bradyarrhythmia, a complication that significantly increases the morbidity and mortality of HF patients and confers an increased risk of sudden cardiac death. Recent studies support the previously unrecognized roles of mitochondrial-sarcoplasmic reticulum connectomics in SAN dysfunction commonly seen with HF. In addition, the roles of distinct AC isoforms that are preferentially expressed and compartmentalized in the SAN to serve a specialized function will be discussed. Finally, the review will consider recent advances in the development of biological pacemakers.
Fibromyalgia (FM) is a chronic pain disorder with a severe impact on a person's health-related quality of life. In addition to the characteristic widespread pain and fatigue, people with FM regularly experience sensory abnormalities and report lingering, often painful sensations after mechanical probing of the skin. The neurobiological changes which underlie the varied symptoms in FM are incompletely understood, but many symptoms and signs can be transferred from patients to mice by administration of patient IgG. The present study aimed to explore whether sensory afferents, including nociceptors, in rodents injected with FM patient IgG displayed ongoing firing after mechanical stimulation. Skin saphenous nerve recordings were retrospectively analyzed to assess the presence and quality of ongoing discharges following mechanical stimulation of sensory afferents in mouse skin after passive-transfer of FM patient IgG. Our analysis revealed that C-mechano and Aδ-mechano afferents innervating the skin in FM-IgG treated animals fired significantly more action potentials after, but not during, mechanical force steps compared to healthy control-IgG treated afferents. This sustained mechanically evoked activity mirrors the nociceptor hyperexcitability observed in people with FM, suggesting that autoreactive IgG might underly this phenomenon in patients. This adds to the growing body of evidence demonstrating that FM symptoms arise from changes in the peripheral nervous system generated by circulating autoreactive IgG. KEY POINTS: Passive transfer of fibromyalgia (FM)-IgG causes sensitization of mechano-sensitive afferents in the skin of mice. Aδ-mechano and C-mechano sensitive afferents in skin from mice injected with FM-IgG display more after-discharge following mechanical stimulation than mice injected with healthy control-IgG. Changes in the peripheral nervous system caused by circulating autoreactive IgG play a role in FM signs and symptoms.