Heartbeats entrain breathing via baroreceptor-mediated modulation of expiratory activity.

NEW FINDINGS: Cardio-ventilatory coupling refers to the onset of inspiration occurring at a preferential latency following the last heartbeat (HB) in expiration. According to the cardiac-trigger hypothesis, the pulse pressure initiates an inspiration via baroreceptor activation. However, the central neural substrate mediating this coupling remains undefined. Using a combination of animal data, human data and mathematical modelling, this study tests the hypothesis that the HB, by way of pulsatile baroreflex activation, controls the initiation of inspiration that occurs through a rapid neural activation loop from the carotid baroreceptors to Bötzinger complex expiratory neurons. ABSTRACT: Cardio-ventilatory coupling refers to a heartbeat (HB) occurring at a preferred latency prior to the next breath. We hypothesized that the pressure pulse generated by a HB activates baroreceptors that modulate brainstem expiratory neuronal activity and delay the initiation of inspiration. In supine male subjects, we recorded ventilation, electrocardiogram and blood pressure during 20-min epochs of baseline, slow-deep breathing and recovery. In in situ rodent preparations, we recorded brainstem activity in response to pulses of perfusion pressure. We applied a well-established respiratory network model to interpret these data. In humans, the latency between a HB and onset of inspiration was consistent across different breathing patterns. In in situ preparations, a transient pressure pulse during expiration activated a subpopulation of expiratory neurons normally active during post-inspiration, thus delaying the next inspiration. In the model, baroreceptor input to post-inspiratory neurons accounted for the effect. These studies are consistent with baroreflex activation modulating respiration through a pauci-synaptic circuit from baroreceptors to onset of inspiration.

Inhibitory control of active expiration by the Bötzinger complex in rats.

KEY POINTS: Contraction of abdominal muscles at the end of expiration during metabolic challenges (such as hypercapnia and hypoxia) improves pulmonary ventilation. The emergence of this active expiratory pattern requires the recruitment of the expiratory oscillator located on the ventral surface of the medulla oblongata. Here we show that an inhibitory circuitry located in the Bötzinger complex is an important source of inhibitory drive to the expiratory oscillator. This circuitry, mediated by GABAergic and glycinergic synapses, provides expiratory inhibition that restrains the expiratory oscillator under resting condition and regulates the formation of abdominal expiratory activity during active expiration. By combining experimental and modelling approaches, we propose the organization and connections within the respiratory network that control the changes in the breathing pattern associated with elevated metabolic demand. ABSTRACT: The expiratory neurons of the Bötzinger complex (BötC) provide inhibitory inputs to the respiratory network, which, during eupnoea, are critically important for respiratory phase transition and duration control. Here, we investigated how the BötC neurons interact with the expiratory oscillator located in the parafacial respiratory group (pFRG) and control the abdominal activity during active expiration. Using the decerebrated, arterially perfused in situ preparations of juvenile rats, we recorded the activity of expiratory neurons and performed pharmacological manipulations of the BötC and pFRG during hypercapnia or after the exposure to short-term sustained hypoxia - conditions that generate active expiration. The experimental data were integrated in a mathematical model to gain new insights into the inhibitory connectome within the respiratory central pattern generator. Our results indicate that the BötC neurons may establish mutual connections with the pFRG, providing expiratory inhibition during the first stage of expiration and receiving excitatory inputs during late expiration. Moreover, we found that application of GABAergic and glycinergic antagonists in the BötC caused opposing effects on abdominal expiratory activity, suggesting complex inhibitory circuitry within the BötC. Using mathematical modelling, we propose that the BötC network organization and its interactions with the pFRG restrain abdominal activity under resting conditions and contribute to abdominal expiratory pattern formation during active expiration observed during hypercapnia or after the exposure to short-term sustained hypoxia.

Cutaneous sensory feedback from paw pads affects lateral balance control during split-belt locomotion in the cat.

Cutaneous sensory feedback from the paw pads plays an important role in regulating body balance, especially in challenging environments like ladder or slope walking. Here, we investigated the contribution of cutaneous sensory feedback from the paw pads to balance control in cats stepping on a split-belt treadmill. Forepaws and hindpaws were anesthetized unilaterally using lidocaine injections. We evaluated body balance in intact and compromised cutaneous feedback conditions during split-belt locomotion with belt-speed ratios of 0.5, 1.0, 1.5 and 2.0. Measures of body balance included step width, relative duration of limb support phases, lateral bias of center of mass (CoM) and margins of static and dynamic stability. In the intact condition, static and dynamic balance declined with increasing belt-speed ratio as a result of a lateral shift of the CoM toward the borders of support on the slower moving belt. Anesthesia of the ipsilateral paws improved locomotor balance with increasing belt-speed ratios by reversing the CoM shift, decreasing the relative duration of the two-limb support phase, increasing the duration of four- or three-limb support phases, and increasing the hindlimb step width and static stability. We observed no changes in most balance measures in anesthetized conditions during tied-belt locomotion at 0.4 m s-1 CoM lateral displacements closely resembled those of the inverted pendulum and of human walking. We propose that unilaterally compromised cutaneous feedback from the paw pads is compensated for by improving lateral balance and by shifting the body toward the anesthetized paws to increase tactile sensation during the stance phase.

Organization of the core respiratory network: Insights from optogenetic and modeling studies.

The circuit organization within the mammalian brainstem respiratory network, specifically within and between the pre-Bötzinger (pre-BötC) and Bötzinger (BötC) complexes, and the roles of these circuits in respiratory pattern generation are continuously debated. We address these issues with a combination of optogenetic experiments and modeling studies. We used transgenic mice expressing channelrhodopsin-2 under the VGAT-promoter to investigate perturbations of respiratory circuit activity by site-specific photostimulation of inhibitory neurons within the pre-BötC or BötC. The stimulation effects were dependent on the intensity and phase of the photostimulation. Specifically: (1) Low intensity (≤ 1.0 mW) pulses delivered to the pre-BötC during inspiration did not terminate activity, whereas stronger stimulations (≥ 2.0 mW) terminated inspiration. (2) When the pre-BötC stimulation ended in or was applied during expiration, rebound activation of inspiration occurred after a fixed latency. (3) Relatively weak sustained stimulation (20 Hz, 0.5-2.0 mW) of pre-BötC inhibitory neurons increased respiratory frequency, while a further increase of stimulus intensity (> 3.0 mW) reduced frequency and finally (≥ 5.0 mW) terminated respiratory oscillations. (4) Single pulses (0.2-5.0 s) applied to the BötC inhibited rhythmic activity for the duration of the stimulation. (5) Sustained stimulation (20 Hz, 0.5-3.0 mW) of the BötC reduced respiratory frequency and finally led to apnea. We have revised our computational model of pre-BötC and BötC microcircuits by incorporating an additional population of post-inspiratory inhibitory neurons in the pre-BötC that interacts with other neurons in the network. This model was able to reproduce the above experimental findings as well as previously published results of optogenetic activation of pre-BötC or BötC neurons obtained by other laboratories. The proposed organization of pre-BötC and BötC circuits leads to testable predictions about their specific roles in respiratory pattern generation and provides important insights into key circuit interactions operating within brainstem respiratory networks.

Interaction between the retrotrapezoid nucleus and the parafacial respiratory group to regulate active expiration and sympathetic activity in rats.

The retrotrapezoid nucleus (RTN) contains chemosensitive cells that distribute CO2-dependent excitatory drive to the respiratory network. This drive facilitates the function of the respiratory central pattern generator (rCPG) and increases sympathetic activity. It is also evidenced that during hypercapnia, the late-expiratory (late-E) oscillator in the parafacial respiratory group (pFRG) is activated and determines the emergence of active expiration. However, it remains unclear the microcircuitry responsible for the distribution of the excitatory signals to the pFRG and the rCPG in conditions of high CO2. Herein, we hypothesized that excitatory inputs from chemosensitive neurons in the RTN are necessary for the activation of late-E neurons in the pFRG. Using the decerebrated in situ rat preparation, we found that lesions of neurokinin-1 receptor-expressing neurons in the RTN region with substance P-saporin conjugate suppressed the late-E activity in abdominal nerves (AbNs) and sympathetic nerves (SNs) and attenuated the increase in phrenic nerve (PN) activity induced by hypercapnia. On the other hand, kynurenic acid (100 mM) injections in the pFRG eliminated the late-E activity in AbN and thoracic SN but did not modify PN response during hypercapnia. Iontophoretic injections of retrograde tracer into the pFRG of adult rats revealed labeled phox2b-expressing neurons within the RTN. Our findings are supported by mathematical modeling of chemosensitive and late-E populations within the RTN and pFRG regions as two separate but interacting populations in a way that the activation of the pFRG late-E neurons during hypercapnia require glutamatergic inputs from the RTN neurons that intrinsically detect changes in CO2/pH.

The Kölliker-Fuse nucleus orchestrates the timing of expiratory abdominal nerve bursting.

Coordination of respiratory pump and valve muscle activity is essential for normal breathing. A hallmark respiratory response to hypercapnia and hypoxia is the emergence of active exhalation, characterized by abdominal muscle pumping during the late one-third of expiration (late-E phase). Late-E abdominal activity during hypercapnia has been attributed to the activation of expiratory neurons located within the parafacial respiratory group (pFRG). However, the mechanisms that control emergence of active exhalation, and its silencing in restful breathing, are not completely understood. We hypothesized that inputs from the Kölliker-Fuse nucleus (KF) control the emergence of late-E activity during hypercapnia. Previously, we reported that reversible inhibition of the KF reduced postinspiratory (post-I) motor output to laryngeal adductor muscles and brought forward the onset of hypercapnia-induced late-E abdominal activity. Here we explored the contribution of the KF for late-E abdominal recruitment during hypercapnia by pharmacologically disinhibiting the KF in in situ decerebrate arterially perfused rat preparations. These data were combined with previous results and incorporated into a computational model of the respiratory central pattern generator. Disinhibition of the KF through local parenchymal microinjections of gabazine (GABAA receptor antagonist) prolonged vagal post-I activity and inhibited late-E abdominal output during hypercapnia. In silico, we reproduced this behavior and predicted a mechanism in which the KF provides excitatory drive to post-I inhibitory neurons, which in turn inhibit late-E neurons of the pFRG. Although the exact mechanism proposed by the model requires testing, our data confirm that the KF modulates the formation of late-E abdominal activity during hypercapnia. NEW & NOTEWORTHY The pons is essential for the formation of the three-phase respiratory pattern, controlling the inspiratory-expiratory phase transition. We provide functional evidence of a novel role for the Kölliker-Fuse nucleus (KF) controlling the emergence of abdominal expiratory bursts during active expiration. A computational model of the respiratory central pattern generator predicts a possible mechanism by which the KF interacts indirectly with the parafacial respiratory group and exerts an inhibitory effect on the expiratory conditional oscillator.

Circadian variability of body temperature responses to methamphetamine.

Vital parameters of living organisms exhibit circadian rhythmicity. Although rats are nocturnal animals, most of the studies involving rats are performed during the day. The objective of this study was to examine the circadian variability of the body temperature responses to methamphetamine. Body temperature was recorded in male Sprague-Dawley rats that received intraperitoneal injections of methamphetamine (Meth, 1 or 5 mg/kg) or saline at 10 AM or at 10 PM. The baseline body temperature at night was 0.8°C higher than during the day. Both during the day and at night, 1 mg/kg of Meth induced monophasic hyperthermia. However, the maximal temperature increase at night was 50% smaller than during the daytime. Injection of 5 mg/kg of Meth during the daytime caused a delayed hyperthermic response. In contrast, the same dose at night produced responses with a tendency toward a decrease of body temperature. Using mathematical modeling, we previously showed that the complex dose dependence of the daytime temperature responses to Meth results from an interplay between inhibitory and excitatory drives. In this study, using our model, we explain the suppression of the hyperthermia in response to Meth at night. First, we found that the baseline activity of the excitatory drive is greater at night. It appears partially saturated and thus is additionally activated by Meth to a lesser extent. Therefore, the excitatory component causes less hyperthermia or becomes overpowered by the inhibitory drive in response to the higher dose. Second, at night the injection of Meth results in reduction of the equilibrium body temperature, leading to gradual cooling counteracting hyperthermia.

Modeling the effects of extracellular potassium on bursting properties in pre-Bötzinger complex neurons.

There are many types of neurons that intrinsically generate rhythmic bursting activity, even when isolated, and these neurons underlie several specific motor behaviors. Rhythmic neurons that drive the inspiratory phase of respiration are located in the medullary pre-Bötzinger Complex (pre-BötC). However, it is not known if their rhythmic bursting is the result of intrinsic mechanisms or synaptic interactions. In many cases, for bursting to occur, the excitability of these neurons needs to be elevated. This excitation is provided in vitro (e.g. in slices), by increasing extracellular potassium concentration (K out) well beyond physiologic levels. Elevated K out shifts the reversal potentials for all potassium currents including the potassium component of leakage to higher values. However, how an increase in K out , and the resultant changes in potassium currents, induce bursting activity, have yet to be established. Moreover, it is not known if the endogenous bursting induced in vitro is representative of neural behavior in vivo. Our modeling study examines the interplay between K out, excitability, and selected currents, as they relate to endogenous rhythmic bursting. Starting with a Hodgkin-Huxley formalization of a pre-BötC neuron, a potassium ion component was incorporated into the leakage current, and model behaviors were investigated at varying concentrations of K out. Our simulations show that endogenous bursting activity, evoked in vitro by elevation of K out , is the result of a specific relationship between the leakage and voltage-dependent, delayed rectifier potassium currents, which may not be observed at physiological levels of extracellular potassium.

Mechanisms of left-right coordination in mammalian locomotor pattern generation circuits: a mathematical modeling view.

The locomotor gait in limbed animals is defined by the left-right leg coordination and locomotor speed. Coordination between left and right neural activities in the spinal cord controlling left and right legs is provided by commissural interneurons (CINs). Several CIN types have been genetically identified, including the excitatory V3 and excitatory and inhibitory V0 types. Recent studies demonstrated that genetic elimination of all V0 CINs caused switching from a normal left-right alternating activity to a left-right synchronized "hopping" pattern. Furthermore, ablation of only the inhibitory V0 CINs (V0D subtype) resulted in a lack of left-right alternation at low locomotor frequencies and retaining this alternation at high frequencies, whereas selective ablation of the excitatory V0 neurons (V0V subtype) maintained the left-right alternation at low frequencies and switched to a hopping pattern at high frequencies. To analyze these findings, we developed a simplified mathematical model of neural circuits consisting of four pacemaker neurons representing left and right, flexor and extensor rhythm-generating centers interacting via commissural pathways representing V3, V0D, and V0V CINs. The locomotor frequency was controlled by a parameter defining the excitation of neurons and commissural pathways mimicking the effects of N-methyl-D-aspartate on locomotor frequency in isolated rodent spinal cord preparations. The model demonstrated a typical left-right alternating pattern under control conditions, switching to a hopping activity at any frequency after removing both V0 connections, a synchronized pattern at low frequencies with alternation at high frequencies after removing only V0D connections, and an alternating pattern at low frequencies with hopping at high frequencies after removing only V0V connections. We used bifurcation theory and fast-slow decomposition methods to analyze network behavior in the above regimes and transitions between them. The model reproduced, and suggested explanation for, a series of experimental phenomena and generated predictions available for experimental testing.

Meth math: modeling temperature responses to methamphetamine.

Methamphetamine (Meth) can evoke extreme hyperthermia, which correlates with neurotoxicity and death in laboratory animals and humans. The objective of this study was to uncover the mechanisms of a complex dose dependence of temperature responses to Meth by mathematical modeling of the neuronal circuitry. On the basis of previous studies, we composed an artificial neural network with the core comprising three sequentially connected nodes: excitatory, medullary, and sympathetic preganglionic neuronal (SPN). Meth directly stimulated the excitatory node, an inhibitory drive targeted the medullary node, and, in high doses, an additional excitatory drive affected the SPN node. All model parameters (weights of connections, sensitivities, and time constants) were subject to fitting experimental time series of temperature responses to 1, 3, 5, and 10 mg/kg Meth. Modeling suggested that the temperature response to the lowest dose of Meth, which caused an immediate and short hyperthermia, involves neuronal excitation at a supramedullary level. The delay in response after the intermediate doses of Meth is a result of neuronal inhibition at the medullary level. Finally, the rapid and robust increase in body temperature induced by the highest dose of Meth involves activation of high-dose excitatory drive. The impairment in the inhibitory mechanism can provoke a life-threatening temperature rise and makes it a plausible cause of fatal hyperthermia in Meth users. We expect that studying putative neuronal sites of Meth action and the neuromediators involved in a detailed model of this system may lead to more effective strategies for prevention and treatment of hyperthermia induced by amphetamine-like stimulants.

Sodium and calcium mechanisms of rhythmic bursting in excitatory neural networks of the pre-Bötzinger complex: a computational modelling study.

The neural mechanisms generating rhythmic bursting activity in the mammalian brainstem, particularly in the pre-Bötzinger complex (pre-BötC), which is involved in respiratory rhythm generation, and in the spinal cord (e.g. locomotor rhythmic activity) that persist after blockade of synaptic inhibition remain poorly understood. Experimental studies in rodent medullary slices containing the pre-BötC identified two mechanisms that could potentially contribute to the generation of rhythmic bursting: one based on the persistent Na(+) current (I(NaP)), and the other involving the voltage-gated Ca(2+) current (I(Ca)) and the Ca(2+) -activated nonspecific cation current (I(CAN)), activated by intracellular Ca(2+) accumulated from extracellular and intracellular sources. However, the involvement and relative roles of these mechanisms in rhythmic bursting are still under debate. In this theoretical/modelling study, we investigated Na(+)-dependent and Ca(2+)-dependent bursting generated in single cells and heterogeneous populations of synaptically interconnected excitatory neurons with I(NaP) and I(Ca) randomly distributed within populations. We analysed the possible roles of network connections, ionotropic and metabotropic synaptic mechanisms, intracellular Ca(2+) release, and the Na(+)/K(+) pump in rhythmic bursting generated under different conditions. We show that a heterogeneous population of excitatory neurons can operate in different oscillatory regimes with bursting dependent on I(NaP) and/or I(CAN), or independent of both. We demonstrate that the operating bursting mechanism may depend on neuronal excitation, synaptic interactions within the network, and the relative expression of particular ionic currents. The existence of multiple oscillatory regimes and their state dependence demonstrated in our models may explain different rhythmic activities observed in the pre-BötC and other brainstem/spinal cord circuits under different experimental conditions.

Towards the Integrative Theory of Alzheimer's Disease: Linking Molecular Mechanisms of Neurotoxicity, Beta-amyloid Biomarkers, and the Diagnosis.

INTRODUCTION: A major gap in amyloid-centric theories of Alzheimer's disease (AD) is that even though amyloid fibrils per se are not toxic in vitro, the diagnosis of AD clearly correlates with the density of beta-amyloid (Aß) deposits. Based on our proposed amyloid degradation toxicity hypothesis, we developed a mathematical model explaining this discrepancy. It suggests that cytotoxicity depends on the cellular uptake of soluble Aß rather than on the presence of amyloid aggregates. The dynamics of soluble beta-amyloid in the cerebrospinal fluid (CSF) and the density of Aß deposits is described using a system of differential equations. In the model, cytotoxic damage is proportional to the cellular uptake of Aß, while the probability of an AD diagnosis is defined by the Aß cytotoxicity accumulated over the duration of the disease. After uptake, Aß is concentrated intralysosomally, promoting the formation of fibrillation seeds inside cells. These seeds cannot be digested and are either accumulated intracellularly or exocytosed. Aß starts aggregating on the extracellular seeds and, therefore, decreases in concentration in the interstitial fluid. The dependence of both Aß toxicity and aggregation on the same process-cellular uptake of Aß-explains the correlation between AD diagnosis and the density of amyloid aggregates in the brain. METHODS: We tested the model using clinical data obtained from the Alzheimer's Disease Neuroimaging Initiative (ADNI), which included records of beta-amyloid concentration in the cerebrospinal fluid (CSF-Aß42) and the density of beta-amyloid deposits measured using positron emission tomography (PET). The model predicts the probability of AD diagnosis as a function of CSF-Aß42 and PET and fits the experimental data at the 95% confidence level. RESULTS: Our study shows that existing clinical data allows for the inference of kinetic parameters describing beta-amyloid turnover and disease progression. Each combination of CSF-Aß42 and PET values can be used to calculate the individual's cellular uptake rate, the effective disease duration, and the accumulated toxicity. We show that natural limitations on these parameters explain the characteristic distribution of the clinical dataset for these two biomarkers in the population. CONCLUSION: The resulting mathematical model interprets the positive correlation between the density of Aß deposits and the probability of an AD diagnosis without assuming any cytotoxicity of the aggregated beta-amyloid. To the best of our knowledge, this model is the first to mechanistically explain the negative correlation between the concentration of Aß42 in the CSF and the probability of an AD diagnosis. Finally, based on the amyloid degradation toxicity hypothesis and the insights provided by mathematical modeling, we propose new pathophysiology-relevant biomarkers to diagnose and predict AD.

Sensory Feedback and Central Neuronal Interactions in Mouse Locomotion.

Locomotion is a complex process involving specific interactions between the central neural controller and the mechanical components of the system. The basic rhythmic activity generated by locomotor circuits in the spinal cord defines rhythmic limb movements and their central coordination. The operation of these circuits is modulated by sensory feedback from the limbs providing information about the state of the limbs and the body. However, the specific role and contribution of central interactions and sensory feedback in the control of locomotor gait and posture remain poorly understood. We use biomechanical data on quadrupedal locomotion in mice and recent findings on the organization of neural interactions within the spinal locomotor circuitry to create and analyze a tractable mathematical model of mouse locomotion. The model includes a simplified mechanical model of the mouse body with four limbs and a central controller composed of four rhythm generators, each operating as a state machine controlling the state of one limb. Feedback signals characterize the load and extension of each limb as well as postural stability (balance). We systematically investigate and compare several model versions and compare their behavior to existing experimental data on mouse locomotion. Our results highlight the specific roles of sensory feedback and some central propriospinal interactions between circuits controlling fore and hind limbs for speed-dependent gait expression. Our models suggest that postural imbalance feedback may be critically involved in the control of swing-to-stance transitions in each limb and the stabilization of walking direction.

Membrane channel hypothesis of lysosomal permeabilization by beta-amyloid.

Alzheimer's disease (AD) is the most common cause of dementia affecting millions of people. Neuronal death in AD is initiated by oligomeric amyloid-ß (Aß) peptides. Recently, we proposed the amyloid degradation toxicity hypothesis, which explains multiple major observations associated with AD including autophagy failure and a decreased metabolism. According to the hypothesis, the key event in the cellular toxicity of amyloid is the formation of non-selective membrane channels in lysosomal membranes by amyloid fragments that are produced by the digestion of Aß previously absorbed by endocytosis. Electrophysiological data suggest that amyloid-formed channels have different sizes, which can be explained by the fact that channel creating barrel-shaped amyloid aggregates can consist of different number of monomers. To estimate the ability of channels to leak molecules of various molecular weights, we modeled the channels as saline-filled cylinders in non-conductive membranes that pass spheres with a density of average globular proteins. As a basis, we used the conductance distribution taken from the previously published experimental dataset, in which single channels with electrical conductance of up to one nanosiemens were registered. Our calculations show that channels with such a giant conductance can allow for passing macromolecules such as large as lysosomal cathepsins implicated in the activation of apoptosis. The formation of giant channels is disproportionally promoted in an acidic environment. Also, amyloid fragments leaking from permeabilized lysosomes can reach the internal leaflet of the plasma membrane and permeabilize it. We conclude that while dissipation of the proton gradient by any (even smallest) amyloid channels readily explains lysosomal failure, the relatively rare events of lysosomal permeabilization to large macromolecules can be an additional mechanism of cellular death induced by exposure to Aß.

Patients with Alzheimer's disease have an increased removal rate of soluble beta-amyloid-42.

Senile plaques, which are mostly composed of beta-amyloid peptide, are the main signature of Alzheimer's disease (AD). Two main forms of beta-amyloid in humans are 40 and 42-amino acid, long; the latter is considered more relevant to AD etiology. The concentration of soluble beta-amyloid-42 (Aß42) in cerebrospinal fluid (CSF-Aß42) and the density of amyloid depositions have a strong negative correlation. However, AD patients have lower CSF-Aß42 levels compared to individuals with normal cognition (NC), even after accounting for this correlation. The goal of this study was to infer deviations of Aß42 metabolism parameters that underlie this difference using data from the Alzheimer's Disease Neuroimaging Initiative cohort. Aß42 is released to the interstitial fluid (ISF) by cells and is removed by several processes. First, growth of insoluble fibrils by aggregation decreases the concentration of soluble beta-amyloid in the ISF. Second, Aß42 is physically transferred from the brain to the CSF and removed with the CSF flow. Finally, there is an intratissue removal of Aß42 ending in proteolysis, which can occur either in the ISF or inside the cells after the peptide is endocytosed. Unlike aggregation, which preserves the peptide in the brain, transfer to the CSF and intratissue proteolysis together represent amyloid removal. Using a kinetic model of Aß42 turnover, we found that compared to NC subjects, AD patients had dramatically increased rates of amyloid removal. A group with late-onset mild cognitive impairment (LMCI) also exhibited a higher rate of amyloid removal; however, this was less pronounced than in the AD group. Estimated parameters in the early-onset MCI group did not differ significantly from those in the NC group. We hypothesize that increased amyloid removal is mediated by Aß42 cellular uptake; this is because CSF flow is not increased in AD patients, while most proteases are intracellular. Aß cytotoxicity depends on both the amount of beta-amyloid internalized by cells and its intracellular conversion into toxic products. We speculate that AD and LMCI are associated with increased cellular amyloid uptake, which leads to faster disease progression. The early-onset MCI (EMCI) patients do not differ from the NC participants in terms of cellular amyloid uptake. Therefore, EMCI may be mediated by the increased production of toxic amyloid metabolites.

Predictions and experimental tests of a new biophysical model of the mammalian respiratory oscillator.

Previously our computational modeling studies (Phillips et al., 2019) proposed that neuronal persistent sodium current (INaP) and calcium-activated non-selective cation current (ICAN) are key biophysical factors that, respectively, generate inspiratory rhythm and burst pattern in the mammalian preBötzinger complex (preBötC) respiratory oscillator isolated in vitro. Here, we experimentally tested and confirmed three predictions of the model from new simulations concerning the roles of INaP and ICAN: (1) INaP and ICAN blockade have opposite effects on the relationship between network excitability and preBötC rhythmic activity; (2) INaP is essential for preBötC rhythmogenesis; and (3) ICAN is essential for generating the amplitude of rhythmic output but not rhythm generation. These predictions were confirmed via optogenetic manipulations of preBötC network excitability during graded INaP or ICAN blockade by pharmacological manipulations in slices in vitro containing the rhythmically active preBötC from the medulla oblongata of neonatal mice. Our results support and advance the hypothesis that INaP and ICAN mechanistically underlie rhythm and inspiratory burst pattern generation, respectively, in the isolated preBötC.

Intermittent hypoxia-induced sensitization of central chemoreceptors contributes to sympathetic nerve activity during late expiration in rats.

Hypertension elicited by chronic intermittent hypoxia (CIH) is associated with elevated activity of the thoracic sympathetic nerve (tSN) that exhibits an enhanced respiratory modulation reflecting a strengthened interaction between respiratory and sympathetic networks within the brain stem. Expiration is a passive process except for special metabolic conditions such as hypercapnia, when it becomes active through phasic excitation of abdominal motor nerves (AbN) in late expiration. An increase in CO(2) evokes late-expiratory (late-E) discharges phase-locked to phrenic bursts with the frequency increasing quantally as hypercapnia increases. In rats exposed to CIH, the late-E discharges synchronized in AbN and tSN emerge in normocapnia. To elucidate the possible neural mechanisms underlying these phenomena, we extended our computational model of the brain stem respiratory network by incorporating a population of presympathetic neurons in the rostral ventrolateral medulla that received inputs from the pons, medullary respiratory compartments, and retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG). Our simulations proposed that CIH conditioning increases the CO(2) sensitivity of RTN/pFRG neurons, causing a reduction in both the CO(2) threshold for emerging the late-E activity in AbN and tSN and the hypocapnic threshold for apnea. Using the in situ rat preparation, we have confirmed that CIH-conditioned rats under normal conditions exhibit synchronized late-E discharges in AbN and tSN similar to those observed in control rats during hypercapnia. Moreover, the hypocapnic threshold for apnea was significantly lowered in CIH-conditioned rats relative to that in control rats. We conclude that CIH may sensitize central chemoreception and that this significantly contributes to the neural impetus for generation of sympathetic activity and hypertension.

Computational modelling of 5-HT receptor-mediated reorganization of the brainstem respiratory network.

Brainstem respiratory neurons express the glycine α(3) receptor (Glyα(3) R), which is a target of modulation by several serotonin (5-HT) receptor agonists. Application of the 5-HT(1A) receptor (5-HT(1A) R) agonist 8-OH-DPAT was shown (i) to depress cellular cAMP, leading to dephosphorylation of Glyα(3) R and augmentation of postsynaptic inhibition of neurons expressing Glyα(3) R (Manzke et al., 2010) and (ii) to hyperpolarize respiratory neurons through 5-HT-activated potassium channels. These processes counteract opioid-induced depression and restore breathing from apnoeas often accompanying pharmacotherapy of pain. The effect is postulated to rely on the enhanced Glyα(3) R-mediated inhibition of inhibitory neurons causing disinhibition of their target neurons. To evaluate this proposal and investigate the neural mechanisms involved, an established computational model of the brainstem respiratory network (Smith et al., 2007), was extended by (i) incorporating distinct subpopulations of inhibitory neurons (glycinergic and GABAergic) and their synaptic interconnections within the Bötzinger and pre-Bötzinger complexes and (ii) assigning the 5-HT(1A) R-Glyα(3) R complex to some of these inhibitory neuron types in the network. The modified model was used to simulate the effects of 8-OH-DPAT on the respiratory pattern and was able to realistically reproduce a number of experimentally observed responses, including the shift in the onset of post-inspiratory activity to inspiration and conversion of the eupnoeic three-phase rhythmic pattern into a two-phase pattern lacking the post-inspiratory phase. The model shows how 5-HT(1A) R activation can produce a disinhibition of inspiratory neurons, leading to the recovery of respiratory rhythm from opioid-induced apnoeas.

Interacting oscillations in neural control of breathing: modeling and qualitative analysis.

In mammalian respiration, late-expiratory (late-E, or pre-inspiratory) oscillations emerge in abdominal motor output with increasing metabolic demands (e.g., during hypercapnia, hypoxia, etc.). These oscillations originate in the retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG) and couple with the respiratory oscillations generated by the interacting neural populations of the Bötzinger (BötC) and pre-Bötzinger (pre-BötC) complexes, representing the kernel of the respiratory central pattern generator. Recently, we analyzed experimental data on the generation of late-E oscillations and proposed a large-scale computational model that simulates the possible interactions between the BötC/pre-BötC and RTN/pFRG oscillations under different conditions. Here we describe a reduced model that maintains the essential features and architecture of the large-scale model, but relies on simplified activity-based descriptions of neural populations. This simplification allowed us to use methods of dynamical systems theory, such as fast-slow decomposition, bifurcation analysis, and phase plane analysis, to elucidate the mechanisms and dynamics of synchronization between the RTN/pFRG and BötC/pre-BötC oscillations. Three physiologically relevant behaviors have been analyzed: emergence and quantal acceleration of late-E oscillations during hypercapnia, transformation of the late-E activity into a biphasic-E activity during hypercapnic hypoxia, and quantal slowing of BötC/pre-BötC oscillations with the reduction of pre-BötC excitability. Each behavior is elicited by gradual changes in excitatory drives or other model parameters, reflecting specific changes in metabolic and/or physiological conditions. Our results provide important theoretical insights into interactions between RTN/pFRG and BötC/pre-BötC oscillations and the role of these interactions in the control of breathing under different metabolic conditions.

Late-expiratory activity: emergence and interactions with the respiratory CpG.

The respiratory rhythm and motor pattern are hypothesized to be generated by a brain stem respiratory network with a rhythmogenic core consisting of neural populations interacting within and between the pre-Bötzinger (pre-BötC) and Bötzinger (BötC) complexes and controlled by drives from other brain stem compartments. Our previous large-scale computational model reproduced the behavior of this network under many different conditions but did not consider neural oscillations that were proposed to emerge within the retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG) and drive preinspiratory (or late-expiratory, late-E) discharges in the abdominal motor output. Here we extend the analysis of our previously published data and consider new data on the generation of abdominal late-E activity as the basis for extending our computational model. The extended model incorporates an additional late-E population in RTN/pFRG, representing a source of late-E oscillatory activity. In the proposed model, under normal metabolic conditions, this RTN/pFRG oscillator is inhibited by BötC/pre-BötC circuits, and the late-E oscillations can be released by either hypercapnia-evoked activation of RTN/pFRG or by hypoxia-dependent suppression of RTN/pFRG inhibition by BötC/pre-BötC. The proposed interactions between BötC/pre-BötC and RTN/pFRG allow the model to reproduce several experimentally observed behaviors, including quantal acceleration of abdominal late-E oscillations with progressive hypercapnia and quantal slowing of phrenic activity with progressive suppression of pre-BötC excitability, as well as to predict a release of late-E oscillations by disinhibition of RTN/pFRG under normal conditions. The extended model proposes mechanistic explanations for the emergence of RTN/pFRG oscillations and their interaction with the brain stem respiratory network.