Unraveling the Mechanisms Underlying Irregularities in Inspiratory Rhythm Generation in a Mouse Model of Parkinson's Disease.

Parkinson's disease (PD) is a neurodegenerative disorder anatomically characterized by a progressive loss of dopaminergic neurons in the substantia nigra compacta (SNpc). Much less known, yet clinically very important, are the detrimental effects on breathing associated with this disease. Consistent with the human pathophysiology, the 6-hydroxydopamine hydrochloride (6-OHDA) rodent model of PD shows reduced respiratory frequency (fR) and NK1r-immunoreactivity in the pre-Bötzinger complex (preBötC) and PHOX2B+ neurons in the retrotrapezoid nucleus (RTN). To unravel mechanisms that underlie bradypnea in PD, we employed a transgenic approach to label or stimulate specific neuron populations in various respiratory-related brainstem regions. PD mice were characterized by a pronounced decreased number of putatively rhythmically active excitatory neurons in the preBötC and adjacent ventral respiratory column (VRC). Specifically, the number of Dbx1 and Vglut2 neurons was reduced by 47.6% and 17.3%, respectively. By contrast, inhibitory Vgat+ neurons in the VRC, as well as neurons in other respiratory-related brainstem regions, showed relatively minimal or no signs of neuronal loss. Consistent with these anatomic observations, optogenetic experiments identified deficits in respiratory function that were specific to manipulations of excitatory (Dbx1/Vglut2) neurons in the preBötC. We conclude that the decreased number of this critical population of respiratory neurons is an important contributor to the development of irregularities in inspiratory rhythm generation in this mouse model of PD.SIGNIFICANCE STATEMENT We found a decreased number of a specific population of medullary neurons which contributes to breathing abnormalities in a mouse model of Parkinson's disease (PD).

Neuroinvasion of SARS-CoV-2 may play a role in the breakdown of the respiratory center of the brain.

The recent outbreak of the novel coronavirus, SARS-CoV-2, has emerged to be highly pathogenic in nature. Although lungs are considered as the primary infected organs by SARS-CoV-2, some of the other organs, including the brain, have also been found to be affected. Here, we have discussed how SARS-CoV-2 might infect the brain. The infection of the respiratory center in the brainstem could be hypothesized to be responsible for the respiratory failure in many COVID-19 patients. The virus might gain entry through the olfactory bulb and invade various parts of the brain, including the brainstem. Alternatively, the entry might also occur from peripheral circulation into the central nervous system by compromising the blood-brain barrier. Finally, yet another possible entry route could be its dispersal from the lungs into the vagus nerve via the pulmonary stretch receptors, eventually reaching the brainstem. Therefore, screening neurological symptoms in COVID-19 patients, especially toward the breakdown of the respiratory center in the brainstem, might help us better understand this disease.

Multi-Level Regulation of Opioid-Induced Respiratory Depression.

Opioids depress minute ventilation primarily by reducing respiratory rate. This results from direct effects on the preBötzinger Complex as well as from depression of the Parabrachial/Kölliker-Fuse Complex, which provides excitatory drive to preBötzinger Complex neurons mediating respiratory phase-switch. Opioids also depress awake drive from the forebrain and chemodrive.

Characterisation of medullary astrocytic populations in respiratory nuclei and alterations in sudden unexpected death in epilepsy.

Central failure of respiration during a seizure is one possible mechanism for sudden unexpected death in epilepsy (SUDEP). Neuroimaging studies indicate volume loss in the medulla in SUDEP and a post mortem study has shown reduction in neuromodulatory neuropeptidergic and monoaminergic neurones in medullary respiratory nuclear groups. Specialised glial cells identified in the medulla are considered essential for normal respiratory regulation including astrocytes with pacemaker properties in the pre-Botzinger complex and populations of subpial and perivascular astrocytes, sensitive to increased pCO2, that excite respiratory neurones. Our aim was to explore niches of medullary astrocytes in SUDEP cases compared to controls. In 48 brainstems from three groups, SUDEP (20), epilepsy controls (10) and non-epilepsy controls (18), sections through the medulla were labelled for GFAP, vimentin and functional markers, astrocytic gap junction protein connexin43 (Cx43) and adenosine A1 receptor (A1R). Regions including the ventro-lateral medulla (VLM; for the pre-Bötzinger complex), Median Raphe (MR) and lateral medullary subpial layer (MSPL) were quantified using image analysis for glial cell populations and compared between groups. Findings included morphologically and regionally distinct vimentin/Cx34-positive glial cells in the VLM and MR in close proximity to neurones. We noted a reduction of vimentin-positive glia in the VLM and MSPL and Cx43 glia in the MR in SUDEP cases compared to control groups (p < 0.05-0.005). In addition, we identified vimentin, Cx43 and A1R positive glial cells in the MSPL region which likely correspond to chemosensory glia identified experimentally. In conclusion, altered medullary glial cell populations could contribute to impaired respiratory regulatory capacity and vulnerability to SUDEP and warrant further investigation.

One bout of neonatal inflammation impairs adult respiratory motor plasticity in male and female rats.

Neonatal inflammation is common and has lasting consequences for adult health. We investigated the lasting effects of a single bout of neonatal inflammation on adult respiratory control in the form of respiratory motor plasticity induced by acute intermittent hypoxia, which likely compensates and stabilizes breathing during injury or disease and has significant therapeutic potential. Lipopolysaccharide-induced inflammation at postnatal day four induced lasting impairments in two distinct pathways to adult respiratory plasticity in male and female rats. Despite a lack of adult pro-inflammatory gene expression or alterations in glial morphology, one mechanistic pathway to plasticity was restored by acute, adult anti-inflammatory treatment, suggesting ongoing inflammatory signaling after neonatal inflammation. An alternative pathway to plasticity was not restored by anti-inflammatory treatment, but was evoked by exogenous adenosine receptor agonism, suggesting upstream impairment, likely astrocytic-dependent. Thus, the respiratory control network is vulnerable to early-life inflammation, limiting respiratory compensation to adult disease or injury.

Impaired chemoreflex correlates with decreased c-Fos in respiratory brainstem centers of the streptozotocin-induced Alzheimer's disease rat model.

Besides impairment in cognition and memory, patients with Alzheimer's disease (AD) often exhibit marked dysfunction in respiratory control. Sleep-disordered breathing (SDB) is commonly found in cases of AD, resulting in periods of hypoxia during sleep. Early structural changes in brainstem areas controlling respiratory function may account for SDB in the course of AD. However, to date the underlying mechanisms for these complications are not known. The streptozotocin (STZ)-induced rat model of AD exhibits abnormal responses to hypoxia and increased astrogliosis in a key region for respiratory control. In this study we further defined the pathophysiological respiratory response of STZ-AD rats to 10% O2. In addition, we analyzed hypoxia-induced neuronal activation in respiratory and cardiovascular nuclei of the dorsal and ventral brainstem. Two hours of hypoxia induced a transient increase in tidal volume that was followed by a prolonged increase in respiratory rate. Only respiratory rate was significantly blunted in the STZ-AD model, which continued over the entire duration of the hypoxic episode. Analysis of c-Fos expression as a marker for neuronal activation showed abundant labeling throughout the nTS, nuclei of the ventral respiratory column, and A1/C1 cells of cardiovascular centers in the ventral brainstem. STZ-AD rats showed a significant decrease of c-Fos labeling in the caudal/medial nTS, rostral ventral respiratory group, and Bötzinger complex. c-Fos in other respiratory centers and A1/C1 cells was unaltered when compared to control. The results of this study document a region-specific impact of STZ-induced AD in respiratory brainstem nuclei. This decrease in c-Fos expression correlates with the observed blunting of respiration to hypoxia in the STZ-AD rat model.

The embryonic development of hindbrain respiratory networks is unaffected by mutation of the planar polarity protein Scribble.

The central command for breathing arises mainly from two interconnected rhythmogenic hindbrain networks, the parafacial respiratory group (pFRG or epF at embryonic stages) and the preBötzinger complex (preBötC), which are comprised of a limited number of neurons located in confined regions of the ventral medulla. In rodents, both networks become active toward the end of gestation but little is known about the signaling pathways involved in their anatomical and functional establishment during embryogenesis. During embryonic development, epF and preBötC neurons migrate from their territories of origin to their final positions in ventral brainstem areas. Planar Cell Polarity (PCP) signaling, including the molecule Scrib, is known to control the developmental migration of several hindbrain neuronal groups. Accordingly, a homozygous mutation of Scrib leads to severe disruption of hindbrain anatomy and function. Here, we aimed to determine whether Scrib is also involved in the prenatal development of the hindbrain nuclei controlling breathing. We combined immunostaining, calcium imaging and electrophysiological recordings of neuronal activity in isolated in vitro preparations. In the Scrib mutant, despite severe neural tube defects, epF and preBötC neurons settled at their expected hindbrain positions. Furthermore, both networks remained capable of generating rhythmically organized, respiratory-related activities and exhibited normal sensitivity to pharmacological agents known to modify respiratory circuit function. Thus Scrib is not required for the proper migration of epF and preBötC neurons during mouse embryogenesis. Our findings thus further illustrate the robustness and specificity of the developmental processes involved in the establishment of hindbrain respiratory circuits.

Supraspinal respiratory plasticity following acute cervical spinal cord injury.

Impaired breathing is a devastating result of high cervical spinal cord injuries (SCI) due to partial or full denervation of phrenic motoneurons, which innervate the diaphragm - a primary muscle of respiration. Consequently, people with cervical level injuries often become dependent on assisted ventilation and are susceptible to secondary complications. However, there is mounting evidence for limited spontaneous recovery of respiratory function following injury, demonstrating the neuroplastic potential of respiratory networks. Although many studies have shown such plasticity at the level of the spinal cord, much less is known about the changes occurring at supraspinal levels post-SCI. The goal of this study was to determine functional reorganization of respiratory neurons in the medulla acutely (>4h) following high cervical SCI. Experiments were conducted in decerebrate, unanesthetized, vagus intact and artificially ventilated rats. In this preparation, spontaneous recovery of ipsilateral phrenic nerve activity was observed within 4 to 6h following an incomplete, C2 hemisection (C2Hx). Electrophysiological mapping of the ventrolateral medulla showed a reorganization of inspiratory and expiratory sites ipsilateral to injury. These changes included i) decreased respiratory activity within the caudal ventral respiratory group (cVRG; location of bulbospinal expiratory neurons); ii) increased proportion of expiratory phase activity within the rostral ventral respiratory group (rVRG; location of inspiratory bulbo-spinal neurons); iii) increased respiratory activity within ventral reticular nuclei, including lateral reticular (LRN) and paragigantocellular (LPGi) nuclei. We conclude that disruption of descending and ascending connections between the medulla and spinal cord leads to immediate functional reorganization within the supraspinal respiratory network, including neurons within the ventral respiratory column and adjacent reticular nuclei.

Minocycline blocks glial cell activation and ventilatory acclimatization to hypoxia.

Ventilatory acclimatization to hypoxia (VAH) is the time-dependent increase in ventilation, which persists upon return to normoxia and involves plasticity in both central nervous system respiratory centers and peripheral chemoreceptors. We investigated the role of glial cells in VAH in male Sprague-Dawley rats using minocycline, an antibiotic that inhibits microglia activation and has anti-inflammatory properties, and barometric pressure plethysmography to measure ventilation. Rats received either minocycline (45mg/kg ip daily) or saline beginning 1 day before and during 7 days of chronic hypoxia (CH, PiO2 = 70 Torr). Minocycline had no effect on normoxic control rats or the hypercapnic ventilatory response in CH rats, but minocycline significantly (P < 0.001) decreased ventilation during acute hypoxia in CH rats. However, minocycline administration during only the last 3 days of CH did not reverse VAH. Microglia and astrocyte activation in the nucleus tractus solitarius was quantified from 30 min to 7 days of CH. Microglia showed an active morphology (shorter and fewer branches) after 1 h of hypoxia and returned to the control state (longer filaments and extensive branching) after 4 h of CH. Astrocytes increased glial fibrillary acidic protein antibody immunofluorescent intensity, indicating activation, at both 4 and 24 h of CH. Minocycline had no effect on glia in normoxia but significantly decreased microglia activation at 1 h of CH and astrocyte activation at 24 h of CH. These results support a role for glial cells, providing an early signal for the induction but not maintenance of neural plasticity underlying ventilatory acclimatization to hypoxia.NEW & NOTEWORTHY The signals for neural plasticity in medullary respiratory centers underlying ventilatory acclimatization to chronic hypoxia are unknown. We show that chronic hypoxia activates microglia and subsequently astrocytes. Minocycline, an antibiotic that blocks microglial activation and has anti-inflammatory properties, also blocks astrocyte activation in respiratory centers during chronic hypoxia and ventilatory acclimatization. However, minocycline cannot reverse ventilatory acclimatization after it is established. Hence, glial cells may provide signals that initiate but do not sustain ventilatory acclimatization.

Catecholaminergic neurons in synaptic connections with pre-Bötzinger complex neurons in the rostral ventrolateral medulla in normoxic and daily acute intermittent hypoxic rats.

The rostral ventrolateral medulla (RVLM) contains cardiovascular-related catecholaminergic neurons and respiratory-related pre-Bötzinger complex (pre-BötC) neurons, which are intermingled and functionally connected for coordinating cardiorespiratory activities. Daily acute intermittent hypoxia (dAIH) is known to elicit respiratory plasticity. However, it is unclear if the catecholaminergic neurons directly synapse onto pre-BötC neurons, and if the local circuitry exhibits plasticity when exposed to dAIH. The present study was aimed to determine the synaptic phenotypes between dopamine-ß-hydroxylase (DßH)-immunoreactive (ir) catecholaminergic neurons and neurokinin 1 receptor (NK1R)-ir pre-BötC neurons, and the effect of dAIH on the neuronal network. Immunofluorescence histochemistry was used to reveal immunoreactivities of DßH and NK1R in the RVLM of normoxic and dAIH rats. Synaptic phenotypes were examined with double-labeling immunoelectron microscopy. We found that DßH immunoreactivity was expressed in somata and processes, some of which were in close apposition to NK1R-ir pre-BötC neurons. DßH-ir gold particles were localized to somata, dendrites, and axonal terminals. DßH-ir terminals formed asymmetric synapses, and occasionally, symmetric synapses in the pre-BötC, featuring the local circuitry. Of the synapses, 28% in normoxic and 29.6% in dAIH groups were apposed to NK1R-ir dendrites. Significant increases in DßH expression and NK1R-ir processes were found in the dAIH group. Moreover, the area and number of processes in close appositions were significantly elevated, strongly suggesting that dAIH induced plasticity with increased connections and interactions between the cardiovascular- and respiratory-related neurons in the local circuitry. In conclusion, asymmetric synapses are predominant in the crosstalk between catecholaminergic and pre-BötC neurons in the RVLM, elaborating excitatory transmission driving the coupling of cardiorespiratory activities. The neural network manifests plasticity in response to dAIH challenge.

Developmental neuropathology of brainstem respiratory centers in unexplained stillbirth: What's the meaning?

Stillbirth is one of the most stressful life events affecting over 3 million pregnancies per year throughout the world. An accurate autopsy of the stillborn fetus, including the placenta and umbilical cord examination, should be performed promptly after delivery. A thorough maternal history also should be taken, including exposures to risk factors. In many cases a death cause, attributable to fetal, maternal, or placental pathology, is clearly identified. However, in 50% or more of cases the cause remains unknown. The purpose of this study is to highlight possible developmental alterations of the autonomic nervous system in unexplained stillbirths to provide an explanation of the pathogenetic mechanism of their death. We conducted a careful neuropathological study of the brainstem, where the main vital centers are located, in 85 unexplained stillbirths and 52 age-matched controls died of known cause. Information on the maternal lifestyle, including the smoking habit, was collected in all cases. Hypodevelopment of neuronal centers involved in breathing control, all connected together in a "respiratory network", precisely hypoplasia of the facial/parafacial complex, Kölliker-Fuse nucleus, pre-Bötzinger nucleus and intermediolateral nucleus, were frequently observed in unexplained deaths, significantly related to maternal cigarette smoking. We support the hypothesis of a strong action of maternal smoking during pregnancy on the development of brainstem respiratory nuclei and suggest an explanation of the high incidence of the respiratory network alterations in unexplained fetal death, when breathing not represents a vital function.

Voltage-Dependent Rhythmogenic Property of Respiratory Pre-Bötzinger Complex Glutamatergic, Dbx1-Derived, and Somatostatin-Expressing Neuron Populations Revealed by Graded Optogenetic Inhibition.

The rhythm of breathing in mammals, originating within the brainstem pre-Bötzinger complex (pre-BötC), is presumed to be generated by glutamatergic neurons, but this has not been directly demonstrated. Additionally, developmental expression of the transcription factor Dbx1 or expression of the neuropeptide somatostatin (Sst), has been proposed as a marker for the rhythmogenic pre-BötC glutamatergic neurons, but it is unknown whether these other two phenotypically defined neuronal populations are functionally equivalent to glutamatergic neurons with regard to rhythm generation. To address these problems, we comparatively investigated, by optogenetic approaches, the roles of pre-BötC glutamatergic, Dbx1-derived, and Sst-expressing neurons in respiratory rhythm generation in neonatal transgenic mouse medullary slices in vitro and also more intact adult perfused brainstem-spinal cord preparations in situ. We established three different triple-transgenic mouse lines with Cre-driven Archaerhodopsin-3 (Arch) expression selectively in glutamatergic, Dbx1-derived, or Sst-expressing neurons for targeted photoinhibition. In each line, we identified subpopulations of rhythmically active, Arch-expressing pre-BötC inspiratory neurons by whole-cell recordings in medullary slice preparations in vitro, and established that Arch-mediated hyperpolarization of these inspiratory neurons was laser power dependent with equal efficacy. By site- and population-specific graded photoinhibition, we then demonstrated that inspiratory frequency was reduced by each population with the same neuronal voltage-dependent frequency control mechanism in each state of the respiratory network examined. We infer that enough of the rhythmogenic pre-BötC glutamatergic neurons also have the Dbx1 and Sst expression phenotypes, and thus all three phenotypes share the same voltage-dependent frequency control property.

Breathing Inhibited When Seizures Spread to the Amygdala and upon Amygdala Stimulation.

Sudden unexpected death in epilepsy (SUDEP) is increasingly recognized as a common and devastating problem. Because impaired breathing is thought to play a critical role in these deaths, we sought to identify forebrain sites underlying seizure-evoked hypoventilation in humans. We took advantage of an extraordinary clinical opportunity to study a research participant with medically intractable epilepsy who had extensive bilateral frontotemporal electrode coverage while breathing was monitored during seizures recorded by intracranial electrodes and mapped by high-resolution brain imaging. We found that central apnea and O2 desaturation occurred when seizures spread to the amygdala. In the same patient, localized electrical stimulation of the amygdala reproduced the apnea and O2 desaturation. Similar effects of amygdala stimulation were observed in two additional subjects, including one without a seizure disorder. The participants were completely unaware of the apnea evoked by stimulation and expressed no dyspnea, despite being awake and vigilant. In contrast, voluntary breath holding of similar duration caused severe dyspnea. These findings suggest a functional connection between the amygdala and medullary respiratory network in humans. Moreover, they suggest that seizure spread to the amygdala may cause loss of spontaneous breathing of which patients are unaware, and thus has potential to contribute to SUDEP. SIGNIFICANCE STATEMENT: Sudden unexpected death in epilepsy (SUDEP) is the most common cause of death in patients with chronic refractory epilepsy. Impaired breathing during and after seizures is common and suspected to play a role in SUDEP. Understanding the cause of this peri-ictal hypoventilation may lead to preventative strategies. In epilepsy patients, we found that seizure invasion of the amygdala co-occurred with apnea and oxygen desaturation, and electrical stimulation of the amygdala reproduced these respiratory findings. Strikingly, the subjects were unaware of the apnea. These findings indicate a functional connection between the amygdala and brainstem respiratory network in humans and suggest that amygdala seizures may cause loss of spontaneous breathing of which patients are unaware-a combination that could be deadly.

Respiratory deficits in a rat model of Parkinson's disease.

Parkinson's disease (PD) is a neurodegenerative disease characterized by loss of the dopaminergic nigrostriatal pathway. In addition to deficits in voluntary movement, PD involves a disturbance of breathing regulation. However, the cause and nature of this disturbance are not well understood. Here, we investigated breathing at rest and in response to hypercapnia (7% CO2) or hypoxia (8% O2), as well as neuroanatomical changes in brainstem regions essential for breathing, in a 6-hydroxydopamine (6-OHDA) rat model of PD. Bilateral injections of 6-OHDA (24µg/µl) into the striatum decreased tyrosine hydroxylase (TH(+))-neurons in the substantia nigra pars compacta (SNpc), transcription factor phox2b-expressing neurons in the retrotrapezoid nucleus and neurokinin-1 receptors in the ventral respiratory column. In 6-OHDA-lesioned rats, respiratory rate was reduced at rest, leading to a reduction in minute ventilation. These animals also showed a reduction in the tachypneic response to hypercapnia, but not to hypoxia challenge. These results suggest that the degeneration of TH(+) neurons in the SNpc leads to impairment of breathing at rest and in hypercapnic conditions. Our data indicate that respiratory deficits in a 6-OHDA rat model of PD are related to downregulation of neural systems involved in respiratory rhythm generation. The present study suggests a new avenue to better understand the respiratory deficits observed in chronic stages of PD.

Respiratory function after selective respiratory motor neuron death from intrapleural CTB-saporin injections.

Amyotrophic lateral sclerosis (ALS) causes progressive motor neuron degeneration, paralysis and death by ventilatory failure. In rodent ALS models: 1) breathing capacity is preserved until late in disease progression despite major respiratory motor neuron death, suggesting unknown forms of compensatory respiratory plasticity; and 2) spinal microglia become activated in association with motor neuron cell death. Here, we report a novel experimental model to study the impact of respiratory motor neuron death on compensatory responses without many complications attendant to spontaneous motor neuron disease. In specific, we used intrapleural injections of cholera toxin B fragment conjugated to saporin (CTB-SAP) to selectively kill motor neurons with access to the pleural space. Motor neuron survival, CD11b labeling (microglia), ventilatory capacity and phrenic motor output were assessed in rats 3-28days after intrapleural injections of: 1) CTB-SAP (25 and 50µg), or 2) unconjugated CTB and SAP (i.e. control; (CTB+SAP). CTB-SAP elicited dose-dependent phrenic and intercostal motor neuron death; 7days post-25µg CTB-SAP, motor neuron survival approximated that in end-stage ALS rats (phrenic: 36±7%; intercostal: 56±10% of controls; n=9; p<0.05). CTB-SAP caused minimal cell death in other brainstem or spinal cord regions. CTB-SAP: 1) increased CD11b fractional area in the phrenic motor nucleus, indicating microglial activation; 2) decreased breathing during maximal chemoreceptor stimulation; and 3) diminished phrenic motor output in anesthetized rats (7days post-25µg, CTB-SAP: 0.3±0.07V; CTB+SAP: 1.5±0.3; n=9; p<0.05). Intrapleural CTB-SAP represents a novel, inducible model of respiratory motor neuron death and provides an opportunity to study compensation for respiratory motor neuron loss.

Drawing breath without the command of effectors: the control of respiration following spinal cord injury.

The maintenance of blood gas and pH homeostasis is essential to life. As such breathing, and the mechanisms which control ventilation, must be tightly regulated yet highly plastic and dynamic. However, injury to the spinal cord prevents the medullary areas which control respiration from connecting to respiratory effectors and feedback mechanisms below the level of the lesion. This trauma typically leads to severe and permanent functional deficits in the respiratory motor system. However, endogenous mechanisms of plasticity occur following spinal cord injury to facilitate respiration and help recover pulmonary ventilation. These mechanisms include the activation of spared or latent pathways, endogenous sprouting or synaptogenesis, and the possible formation of new respiratory control centres. Acting in combination, these processes provide a means to facilitate respiratory support following spinal cord trauma. However, they are by no means sufficient to return pulmonary function to pre-injury levels. A major challenge in the study of spinal cord injury is to understand and enhance the systems of endogenous plasticity which arise to facilitate respiration to mediate effective treatments for pulmonary dysfunction.

Laser ablation of Dbx1 neurons in the pre-Bötzinger complex stops inspiratory rhythm and impairs output in neonatal mice.

To understand the neural origins of rhythmic behavior one must characterize the central pattern generator circuit and quantify the population size needed to sustain functionality. Breathing-related interneurons of the brainstem pre-Bötzinger complex (preBötC) that putatively comprise the core respiratory rhythm generator in mammals are derived from Dbx1-expressing precursors. Here, we show that selective photonic destruction of Dbx1 preBötC neurons in neonatal mouse slices impairs respiratory rhythm but surprisingly also the magnitude of motor output; respiratory hypoglossal nerve discharge decreased and its frequency steadily diminished until rhythm stopped irreversibly after 85±20 (mean ± SEM) cellular ablations, which corresponds to ∼15% of the estimated population. These results demonstrate that a single canonical interneuron class generates respiratory rhythm and contributes in a premotor capacity, whereas these functions are normally attributed to discrete populations. We also establish quantitative cellular parameters that govern network viability, which may have ramifications for respiratory pathology in disease states.

Sudden unexpected death owing to unilateral medial medullary infarction with early involvement of the respiratory center.

A 64-year-old woman was found dead in her home. At autopsy, although relatively fresh bruises were found on her body, no lethal injury was observed in an internal observation. Mild edematous swelling of the right half of the medulla oblongata was observed. There was acute medial medullary infarction (MMI), which mainly involved the nucleus hypoglossi, medial lemniscus, hypoglossal root, inferior olivary nucleus, and pyramidal tract. Subacute infarction of the lower part of the cerebellum was also found, and severe atherosclerosis of the right vertebral artery containing thrombi was found as the culprit lesion. Immunohistochemistry using amyloid precursor protein (APP) was positive in neuronal tissue in the nucleus ambiguus, despite not showing coagulative necrosis in the nucleus. Therefore, acute ischemic necrosis of the nucleus ambiguus, which is considered to be a component of the dorsal respiratory group, may be a significant finding for her expected death. Immunohistochemistry of APP may be useful for confirming the precise extent of acute ischemia in brain stem infarction, such as unilateral MMI.

Parabrachial nucleus involvement in multiple system atrophy.

UNLABELLED: Multiple system atrophy (MSA) is associated with respiratory dysfunction, including sleep apnea, respiratory dysrhythmia, and laryngeal stridor. Neurons of the parabrachial nucleus (PBN) control respiratory rhythmogenesis and airway resistance. OBJECTIVES: The objective of this study is to determine whether there was involvement of putative respiratory regions of the PBN in MSA. METHODS: We examined the pons at autopsy in 10 cases with neuropathologically confirmed MSA and 8 age-matched controls. Sections obtained throughout the pons were processed for calcitonin-gene related peptide (CGRP) and Nissl staining to identify the lateral crescent of the lateral PBN (LPB) and the Kölliker-Fuse nucleus (K-F), which are involved in respiratory control. Cell counts were performed using stereology. RESULTS: There was loss of CGRP neurons in the PBN in MSA (total estimated cell counts for the external LPB cluster was 12,584 ± 1146 in controls and 5917 ± 389 in MSA, p<0.0001); for the external medial PBN (MPB) cluster it was 15,081 ± 1758 in controls and 7842 ± 466 in MSA, p<0.001. There was also neuronal loss in putative respiratory regions of the PBN, including the lateral crescent of the LPB (13,039 ± 1326 in controls and 4164 ± 872 in MSA, p<0.0001); and K-F (5120 ± 495 in controls and 999 ± 308 in MSA, p<0.0001). CONCLUSIONS: There is involvement of both CGRP and putative respiratory cell groups in the PBN in MSA. Whereas the clinical implications of CGRP cell loss are still undetermined, involvement of the LPB and K-F may contribute to respiratory dysfunction in this disorder.

[Damage effects of chronic hypoxia on medulla oblongata associated with oxidative stress and cell apoptosis].

The aim of this study is to study the damage effects of chronic hypoxia on medulla oblongata and to explore whether the damage is associated with oxidative stress and cell apoptosis. Adult male SD rats were randomly divided into two groups: control group and chronic hypoxia group. Medulla oblongata was obtained for the following methods of analyses. Nissl's staining was used to examine the Niss bodies of neurons in medullary respiratory related nuclei, biochemistry methods were utilized to examine oxidant stress damage induced by chronic hypoxia on medulla oblongata through measuring malondialdehyde (MDA) content and superoxide dismutase (SOD) activity, and RT-PCR technique was used to study the influence of apoptosis induced by chronic hypoxia on medulla oblongata through analyzing the levels of Bax mRNA and Bcl-2 mRNA. The results showed the optical densities of Nissl's staining in pre-BötC, NA, NTS, FN, and 12N were significantly decreased in chronic hypoxia group in comparison with that in control group (P < 0.05). In chronic hypoxia group, MDA level was significantly higher than that in the control group (P < 0.05), whereas SOD level had no significant difference between the two groups (P > 0.05). Bax mRNA expression had no obvious change and Bcl-2 mRNA expression significantly decreased in chronic hypoxia group in comparison with that in control group (P < 0.05). The results suggest that chronic hypoxia could bring about serious damage to medullary respiratory centers through aggravating oxidative stress and increasing cell apoptosis.