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The carotid body (CB) is the key oxygen sensing organ. While the expression of CB specific genes is relatively well studied in animals, corresponding data for the human CB are missing. In this study we used five surgically removed human CBs to characterize the CB transcriptome with microarray and PCR analyses, and compared the results with mice data. In silico approaches demonstrated a unique gene expression profile of the human and mouse CB transcriptomes and an unexpected upregulation of both human and mouse CB genes involved in the inflammatory response compared to brain and adrenal gland data. Human CBs express most of the genes previously proposed to be involved in oxygen sensing and signalling based on animal studies, including NOX2, AMPK, CSE and oxygen sensitive K+ channels. In the TASK subfamily of K+ channels, TASK-1 is expressed in human CBs, while TASK-3 and TASK-5 are absent, although we demonstrated both TASK-1 and TASK-3 in one of the mouse reference strains. Maxi-K was expressed exclusively as the spliced variant ZERO in the human CB. In summary, the human CB transcriptome shares important features with the mouse CB, but also differs significantly in the expression of a number of CB chemosensory genes. This study provides key information for future functional investigations on the human carotid body.

We have previously shown that the adult DBA/2J and A/J strains of mice differ in carotid body volume and morphology. The question has arisen whether these differences develop during the prenatal or postnatal period. Investigating morphological development of the carotid body and contributing genes in these mice can provide further understanding of the appropriate formation of the carotid body. We examined the carotid body of these mice from 1 day to 4 wk old for differences in volume, morphology, and gene expression of Gdnf family, Dlx2, Msx2, and Phox2b. The two strains showed divergent morphology starting at 1 wk old. The volume of the carotid body increased from 1 wk up to 2 wk old to the level of 4 wk old in the DBA/2J mice but not in the A/J mice. This corresponds with immunoreactivity of LC3, an autophagy marker, in A/J tissues at 10 days and 2 wk. The differences in gene expression were examined at 1 wk, 10 days, and 2 wk old, because divergent growth occurred during this period. The DBA/2J's carotid body at 1 wk old showed a greater expression of Msx2 than the A/J's carotid body. No other candidate genes showed consistent differences between the ages and strains. The difference was not seen in sympathetic cervical ganglia of 1 wk old, suggesting that the difference is carotid body specific. The current study indicates the critical postnatal period for developing distinctive morphology of the carotid body in these mice. Further studies are required to further elucidate a role of Msx2 and other uninvestigated genes.

Acetylcholine (ACh) has been considered an important excitatory neurotransmitter in the carotid body (CB). Its physiological and pharmacological effects, metabolism, release, and receptors have been well documented in several species. Various nicotinic and muscarinic ACh receptors are present in both afferent nerve endings and glomus cells. Therefore, ACh can depolarize or hyperpolarize the cell membrane depending on the available receptor type in the vicinity. Binding of ACh to its receptor can create a wide variety of cellular responses including opening cation channels (nicotinic ACh receptor activation), releasing Ca(2+) from intracellular storage sites (via muscarinic ACh receptors), and modulating activities of K(+) and Ca(2+) channels. Interactions between ACh and other neurotransmitters (dopamine, adenosine, nitric oxide) have been known, and they may induce complicated responses. Cholinergic biology in the CB differs among species and even within the same species due to different genetic composition. Development and environment influence cholinergic biology. We discuss these issues in light of current knowledge of neuroscience.

The carotid body (CB) is the primary hypoxic chemosensory organ. Its hypoxic response appears to be genetically controlled. We have hypothesized that: 1) genes related to CB function are expressed less in the A/J mice (low responder to hypoxia) compared with DBA/2J mice (high responder to hypoxia); and 2) gene expression levels of morphogenic and trophic factors of the CB are significantly lower in the A/J mice than DBA/2J mice. This study utilizes microarray analysis to test these hypotheses. Three sets of CBs were harvested from both strains. RNA was isolated and used for global gene expression profiling (Affymetrix Mouse 430 v2.0 array). Statistically significant gene expression was determined as a minimum six counts of nine pairwise comparisons, a minimum 1.5-fold change, and P ≤ 0.05. Our results demonstrated that 793 genes were expressed less and that 568 genes were expressed more in the A/J strain vs. the DBA/2J strain. Analysis of individual genes indicates that genes encoding ion channels are differentially expressed between the two strains. Genes related to neurotransmitter metabolism, synaptic vesicles, and the development of neural crest-derived cells are expressed less in the A/J CB vs. the DBA/2J CB. Through pathway analysis, we have constructed a model that shows gene interactions and offers a roadmap to investigate CB development and hypoxic chemosensing/chemotransduction processes. Particularly, Gdnf, Bmp2, Kcnmb2, Tph1, Hif1a, and Arnt2 may contribute to the functional differences in the CB between the two strains. Bmp2, Phox2b, Dlx2, and Msx2 may be important for the morphological differences.

In a previous study, DBA/2J and A/J inbred mice showed extremely different hypoxic ventilatory responses, suggesting variations in their carotid bodies. We have assessed the morphological and functional differences of the carotid bodies in these mice. Histological examination revealed a clearly delineated carotid body only in the DBA/2J mice. Many typical glomus cells and glomeruli appeared in the DBA/2J but not in the A/J mice. The size of the carotid body in the DBA/2J and A/J mice was 6.3 ± 0.5 × 106and 1.5 ± 0.3 × 106μm3, respectively. The area immunostained for tyrosine hydroxylase, an estimation of the glomus cell quantity, was four times larger in the DBA/2J mice than in the A/J mice. The individual data points in the DBA/2J mice segregated from those in the A/J mice. ACh increased intracellular Ca2+in most clusters (81%) of cultured carotid body cells from the DBA/2J mice, but only in 18% of clusters in the A/J mice. These data suggest that genetic determinants account for the strain differences in the structure and function of the carotid body.

Traumatic brain injury (TBI) causes severe disruption of cerebral electrical activity and electroencephalography (EEG) is emerging as a standard tool to monitor TBI patients in the acute period of risk for secondary injuries. However, animal studies of EEG pathology in the context of TBI are surprisingly sparse, largely because of the lack of real-time continuous EEG (cEEG) monitoring in animal TBI models. Here, we performed long-term EEG monitoring to study nonconvulsive seizures (NCS), periodic epileptiform discharges (PED), and EEG power spectra following three injury severity levels in a rat model of penetrating ballistic-like brain injury (PBBI). EEG signals were recorded continuously from bilateral hemispheres of freely behaving rats for 72 h and for 2 h on days 7 and 14 after the injury. We report that the incidence of NCS and PED positively correlated with the injury severity, where 13%, 39%, and 59% of the animals exhibited NCS, and 0%, 30%, and 65% of the animals exhibited PED following 5%, 10% and 12.5% PBBI, respectively. Similar correlations existed for the number of NCS and PED events and their duration. NCS and PED occurred either independently or in tandem. Longer NCS durations were associated with larger lesion volumes. Significant EEG slowing evidenced by the EEG power shift toward the δ frequency band (0.5-4 Hz) occurred within 2 h after PBBI, which resolved over time but persisted longer after greater injury severity. In contrast, decreases in higher frequency power (i.e., 30-35 Hz) remained depressed throughout 14 days. This is the first long-term cEEG study of the acute injury phase in a rat model of severe TBI, demonstrating common occurrences of clinically observed electrocortical pathology, such as NCS, PED, and cortical slowing. These EEG pathologies may serve as critical care biomarkers of brain injury, and offer clinically relevant metrics for studying acute therapeutic interventions.

The ventilatory response to hypoxia depends on the carotid body function and sleep-wake states. Therefore, the response must be measured in a consistent sleep-wake state. In mice, EMG with behavioral indices (coordinated movements, CMs; myoclonic twitches, MTs) has been used to assess sleep-wake states. However, in neonatal mice EMG instrumentation could induce stress, altering their behavior and ventilation. Accordingly, we examined: (1) if EMG can be eliminated for assessing sleep-wake states; and (2) behavioral characteristics and carotid body-mediated respiratory control during sleep with EMG (EMG+) or without EMG (EMG-). Seven-day-old DBA/2J and A/J mice were divided into EMG+ and EMG- groups. In both strains, CMs occurred when EMG was high; MTs were present during silent/low EMG activity. The durations of high EMG activity and of CMs were statistically indifferent. Thus, CMs can be used to indicate wake state without EMG. The stress caused by EMG instrumentation may be distinctively manifested based on genetic background. Prolonged agitation was observed in some EMG+ DBA/2J (5 of 13), but not in A/J mice. The sleep time and MT counts were indifferent between the groups in DBA/2J mice. The EMG+ A/J group showed longer sleep time and less MT counts than the EMG- A/J group. Mean respiratory variables (baseline, hyperoxic/hypoxic responses) were not severely influenced by EMG+ in either strain. Individual values were more variable in EMG+ mice. Carotid body-mediated respiratory responses (decreased ventilation upon hyperoxia and increased ventilation upon mild hypoxia) during sleep were clearly observed in these neonatal mice with or without EMG instrumentation.

This brief review first touches on the origins of the earth's oxygen. It then identifies and locates the principal oxygen sensor in vertebrates, the carotid body (CB). The CB is unique in that in human subjects, it is the only sensor of lower than normal levels in the partial pressure of oxygen (hypoxia, HH). Another oxygen sensor, the aortic bodies, are mostly vestigial in higher vertebrates. At least they play a much smaller role than the CB. In such an important role, the many reflexes in response to CB stimulation by HH are presented. After briefly reviewing what CB stimulation does, the next topic is to describe how the CB chemotransduces HH into neural signals to the brain. Several mechanisms are known, but critical steps in the mechanisms of chemosensation and chemotransduction are still under investigation. Finally, a brief glance at the operation of the CB in chronic heart failure patients is presented. Specifically, the role of nitric oxide, NO, is discussed.

The Carotid Body (CB) senses hypoxia, hypercapnia, and acidosis in the arterial blood. The resulting increase in CB neural output (CBNO) to the nucleus tractus solitarius in the medulla promotes reflex responses in the respiratory, circulatory, renal, and endocrine systems. Increases in CBNO are commonly thought to be due to the release of neurotransmitters from glomus cells in the CB. Additional to the action of these released transmitters on the postsynaptic afferent neurons which abut on the glomus cells the transmitters act presynaptically on glomus cell autoreceptors. Among the several transmitters contained in the glomus cells there now exists considerable evidence supporting excitatory roles for both acetylcholine (ACh) and ATP and an inhibitory role for dopamine (DA) and norepinephrine (NE) (Fitzgerald, 2000). The release of ACh (Fitzgerald et al., 1999; Kim et al., 2004) and catecholamines (Wang and Fitzgerald, 2002) appears to be influenced by modulators. The present study investigated the action of adenosine (ADO) on the release of ACh, DA, and NE since it has been reported that ADO influences CBNO (McQueen and Ribeiro, 1981) and CB-mediated increases in ventilation (Monteiro and Ribeiro, 1987). The study further investigated the action of nitric oxide (NO) on the release of ACh since NO has been reported to reduce the hypoxia-induced increase in CBNO (Wang, et al., 1994).

The role of the carotid body (CB) in response to hypoxia is very well defined (Fitzgerald and Shirahata, 1997). The hypoxic ventilatory response (HVR) is characterized by an increase in ventilation, but this response remains variable among individuals (Eisele et al., 1992; Vizek et al., 1987; Weil 1970). Genetics may play a critical role in explaining this variability. Indeed, longitudinal and twin studies do demonstrate the role of genetics in the HVR (Collins et al., 1978; Kawakami et al., 1982). Studies utilizing inbred strains of mice have also demonstrated the effect of genetics on the response to hypoxia (Tankersley et al., 1994 & 2000). Two strains of mice in these studies were identified as having extreme responses to hypoxia. The DBA/2J strain demonstrated the highest HVR, whereas the A/J strain demonstrated the lowest HVR (Tankersley et al., 1994). In another study analyzing the role of genetics in a mouse model of sleepinduced hypoxia, the DBA/2J strain demonstrated an increased sensitivity to hypoxia during sleep, compared to that of the A/J strain (Rubin et al., 2004). In order to elucidate a potential explanation that may contribute to this difference in hypoxic sensitivity, we examined CB morphology and volume in adult DBA/2J and A/J strains (Yamaguchi et al., 2003). Results demonstrated a significantly larger volume as well as an increased glomus cell quantity in the CB of DBA/2J strain compared to that of the A/J strain. A question arises whether these differences exist from early neonatal ages. Or, developmental plasticity may contribute to these strain differences. In this study, we analyzed CB volume, glomus cell quantity, and ventilation during development in the DBA/2J and A/J strains of mice. We also introduced a glimpse into the genetic influence on chemoreceptor development with emphasis on the role of glial-cell-line-derived neurotrophic factor (GDNF).

The carotid body is a major chemosensory organ for hypoxia, hypercapnia and acidosis in the arterial blood (Fitzgerald and Shirahata 1997; Gonzalez et al., 1994). During hypoxia, the neural output from the carotid body increases and reflexly modifies several variables in the respiratory system. A prominent response is an increase in ventilation, but the hypoxic ventilatory response (HVR) among individuals varies widely (Eisele et al., 1992; Vizek et al., 1987). Studies in humans (Collins et al., 1978; Kawakami et al., 1982; Nishimura et al., 1991; Thomas et al., 1993) suggest that genetic factors significantly contribute to those differences. Similarly, studies in mice (Tankersley et al., 1994) and rats (Weil et al., 1998) clearly indicate that genetic determinants robustly influenced HVR. Among several inbred strains of mice the DBA/2J mice demonstrated the highest HVR and the A/J mice the lowest HVR (Tankersley et al., 1994). The differences in HVR between these two strains of mice may be closely related to the structural differences of the carotid body (Yamaguchi et al., 2003). The size of the carotid body and the quantity of glomus cells in the DBA/2J mice are significantly larger than those in the A/J mice. Those differences were clearly segregated between the strains, suggesting that genetic factors strongly influence the observed phenotypic differences between the DBA/2J and A/J mice. The purpose of the current study was to confirm that the morphological characteristic differences in the carotid body between the DBA/2J and A/J mice are controlled by genetic factors. Thus, we generated the first-filial progeny (F1) by a crossing the DBA/2J (female) and A/J (male) strains of mice, and examined the morphological differences of the carotid body in the DBA/2J, A/J and their F1 mice.

The impact of genetic variation on cardiovascular responses to hypoxia and hypercapnia is not well understood. Therefore, we determined the acute changes in systemic arterial blood pressure (PSA) and heart rate (HR) in seven strains of commonly used inbred mice exposed to acute periods of hypoxia (10% O2), hypercapnia (5% CO2), and hypoxia/hypercapnia (10% O2 + 5% CO2) during wakefulness. Hypercapnia induced an essentially homogeneous response across strains, with PSA maintained at or slightly above baseline and with HR exhibiting a typical baroreceptor-mediated bradycardia. In contrast, exposure to hypoxia elicited a marked heterogeneity in cardiovascular responses between strains. The change in PSA during hypoxia ranged from maintenance of normotension in the FVB/J strain to profound hypotension of ∼30 mmHg in the DBA/2J strain. HR responses were highly variable between strains during hypoxia, and with the exception of the DBA/2J strain that exhibited significant bradyarrhythmias and consequent hypotension, the HR responses were unrelated to changes in PSA. The PSA response to combined hypoxia/hypercapnia represented a balance of the hypertension of hypercapnia and the hypotension of hypoxia in six of the seven strains. In the FVB/J strain, combined hypoxia/hypercapnia produced a hypertensive response that was greater than that of hypercapnia alone. These results suggest that genetic background affects the cardiovascular response to hypoxia, but not hypercapnia.

The carotid body is a major arterial chemoreceptor that senses low O2 tension, high CO2 tension and low pH in the arterial blood. It is generally believed that neurotransmitters, including acetylcholine (ACh), participate in the genesis of afferent neural output from the carotid body and modulate the function of chemoreceptor cells (glomus cells). Previous pharmacological studies suggest that M1 and M2 muscarinic ACh receptors (mAChRs) are involved in these processes. This study was designed to demonstrate the presence and localization of M1 and M2 mAChRs in the carotid body and in the petrosal ganglion of the cat. Since DNA sequences of the cat M1 and M2 mAChRs were not known, we first determined partial DNA sequences. These sequences and deduced amino acid sequences highly resembled those of human and the rat. Subsequent reverse transcription-polymerase chain reaction (RT-PCR)analysis has demonstrated that mRNAs for M1 and M2 mAChRs are present in the carotid body and the petrosal ganglion of the cat. Immunohistochemistry has indicated that the localization of these receptors appears different. Immunoreactivity for M1 mAChR was strong in nerves in the carotid body. Nerve endings positively stained for M1 mAChR appear to innervate glomus cells. Weak staining for M1 mAChRs was seen in glomus cells. On the other hand, M2 receptor protein seems to be present in glomus cells but not on nerve endings. One third of the neurons in the petrosal ganglion showed immunoreactivity for M1 mAChR. Many neurons and nerve fibers in the petrosal ganglion expressed M2 mAChR immunoreactivity. The results were consistent with previous pharmacological studies. Thus, activation of M1 mAChRs on afferent nerve endings may be linked to the increase in neural output during hypoxia. Further, M1 and M2 mAChRs on glomus cells modulate the release of neurotransmitters.

Exogenously administered adenosine provokes an increase in respiration in both animal models and in man. Administered near the carotid body adenosine increases neural output from the carotid body in rats and cats. Hypoxia has the same effect. Hypoxia also provokes a release of acetylcholine (ACh), dopamine (DA), and norepinephrine (NE) from the carotid body. The present study aimed to determine the effect of exogenous adenosine on the release of ACh, DA, and NE from the carotid bodies of cats. After a recovery period (from surgery) carotid bodies were first incubated for 10 (DA, NE) or 15 (ACh) min in Eppendorf tubes containing 85 μL of a physiological salt solution equilibrated with 40% O 2 /5% CO 2 at 37 °C (hyperoxia). At the end of the incubation period the medium was drawn off, and measured for ACh, DA, and NE using HPLC-ECD methods. Next 85 μL of the medium and the tubes were equilibrated with a hypoxic gas mixture (4% O 2 /5% CO 2 ) and the carotid bodies were incubated for 10 (DA, NE) or 15 (ACh) min, at the end of which the medium was drawn off and measured for ACh, DA, and NE. In the ACh studies there followed a post-hypoxic hyperoxic exposure (40% O 2 /5% CO 2 ). ACh tubes were then made 100 μM with respect to adenosine, and the hyperoxic, hypoxic, and post-hypoxic hyperoxic challenges were repeated. One of the two DA, NE tubes had the 100 μM adenosine from the start. Adenosine significantly increased the release of ACh, but significantly decreased the hypoxia-induced release of DA. Potential mechanisms for these changes are reviewed.

In obstructive sleep apnea, hypoxic ventilatory sensitivity may affect the degree of hypoxic stress and sleep disruption that occurs in response to upper airway obstruction. We induced (1) sleep-induced hypoxia (SIH) or (2) sleep fragmentation (SF) without hypoxia for 5 days (12-hour light/dark cycle) in two inbred mouse strains with low (A/J) and high (DBA/2J) hypoxic ventilatory sensitivities. During SIH, the time to arousal (26.4 +/- 1.1 vs. 21.3 +/- 1.5 seconds, p0.025) and the severity of hypoxic exposure (nadir FIO2: 11.5 +/- 0.4 vs. 13.6 +/- 0.1%, p0.002) was greater in A/J than DBA/2J mice. Furthermore, A/J mice had a greater frequency of hypoxic events (640 +/- 29 vs. 368 +/- 33 events per 24 hours, p0.001) and total sleep time (47.5 +/- 2.8% vs. 26.5 +/- 2.4% per 24 hours, p0.0001) during SIH than DBA/2J mice. In contrast, the event characteristics and total sleep time during SF were the same in both strains. Furthermore, in the light phase, both strains showed a longer (p0.01) time to arousal during SIH and SF compared with the dark phase. We conclude that genetic background can influence respiratory events and sleep architecture during SIH and that the arousal threshold is subject to circadian variation. Our data imply that individuals with low hypoxic sensitivity may be at a greater risk for hypoxia-related complications of obstructive sleep apnea.

To investigate the pathophysiological sequelae of sleep-disordered breathing (SDB), we have developed a mouse model in which hypoxia was induced during periods of sleep and was removed in response to arousal or wakefulness. An on-line sleep-wake detection system, based on the frequency and amplitude of electroencephalograph and electromyograph recordings, served to trigger intermittent hypoxia during periods of sleep. In adult male C57BL/6J mice ( n= 5), the sleep-wake detection system accurately assessed wakefulness (97.2 ± 1.1%), non-rapid eye movement (NREM) sleep (96.0 ± 0.9%) and rapid eye movement (REM) sleep (85.6 ± 5.0%). After 5 consecutive days of SDB, 554 ± 29 (SE) hypoxic events were recorded over a 24-h period at a rate of 63.6 ± 2.6 events/h of sleep and with a duration of 28.2 ± 0.7 s. The mean nadir of fraction of inspired O2 (Fi O2 ) on day 5 was 13.2 ± 0.1%, and 137.1 ± 13.2 of the events had a nadir Fi O2 2. Arterial blood gases confirmed that hypoxia of this magnitude lead to a significant degree of hypoxemia. Furthermore, 5 days of SDB were associated with decreases in both NREM and REM sleep during the light phase compared with the 24-h postintervention period. We conclude that our murine model of SDB mimics the rate and magnitude of sleep-induced hypoxia, sleep fragmentation, and reduction in total sleep time found in patients with moderate to severe SDB in the clinical setting.