Characteristics of carotid body chemosensitivity in NADPH oxidase-deficient mice

2002 ◽  
Vol 282 (1) ◽  
pp. C27-C33 ◽  
Author(s):  
L. He ◽  
J. Chen ◽  
B. Dinger ◽  
K. Sanders ◽  
K. Sundar ◽  
...  
Keyword(s):  
Tyrosine Hydroxylase ◽  
Nadph Oxidase ◽  
Carotid Body ◽  
Gene Knockout ◽  
Type I ◽  
Type I Cells ◽  
Carotid Bodies ◽  
Marker Antigen ◽  
I Cells

Various heme-containing proteins have been proposed as primary molecular O2 sensors for hypoxia-sensitive type I cells in the mammalian carotid body. One set of data in particular supports the involvement of a cytochrome b NADPH oxidase that is commonly found in neutrophils. Subunits of this enzyme have been immunocytochemically localized in type I cells, and diphenyleneiodonium, an inhibitor of the oxidase, increases carotid body chemoreceptor activity. The present study evaluated immunocytochemical and functional properties of carotid bodies from normal mice and from mice with a disrupted gp91 phagocytic oxidase (gp91 phox ) DNA sequence gene knockout (KO), a gene that codes for a subunit of the neutrophilic form of NADPH oxidase. Immunostaining for tyrosine hydroxylase, a signature marker antigen for type I cells, was found in groups or lobules of cells displaying morphological features typical of the O2-sensitive cells in other species, and the incidence of tyrosine hydroxylase-immunopositive cells was similar in carotid bodies from both strains of mice. Studies of whole cell K+currents also revealed identical current-voltage relationships and current depression by hypoxia in type I cells dissociated from normal vs. KO animals. Likewise, hypoxia-evoked increases in intracellular Ca2+ concentration were not significantly different for normal and KO type I cells. The whole organ response to hypoxia was evaluated in recordings of carotid sinus nerve activity in vitro. In these experiments, responses elicited by hypoxia and by the classic chemoreceptor stimulant nicotine were also indistinguishable in normal vs. KO preparations. Our data demonstrate that carotid body function remains intact after sequence disruption of the gp91 phox gene. These findings are not in accord with the hypothesis that the phagocytic form of NADPH oxidase acts as a primary O2 sensor in arterial chemoreception.

scholarly journals Immunohistochemical Characteristics of the Human Carotid Body in the Antenatal and Postnatal Periods of Development

2021 ◽  
Vol 22 (15) ◽  
pp. 8222
Author(s):  
Dmitry Otlyga ◽  
Ekaterina Tsvetkova ◽  
Olga Junemann ◽  
Sergey Saveliev
Keyword(s):  
Tyrosine Hydroxylase ◽  
Carotid Body ◽  
Nerve Fibers ◽  
Individual Development ◽  
Endocrine Function ◽  
Type I ◽  
Type I Cells ◽  
Carotid Bodies ◽  
I Cells ◽  
Human Carotid

The evolutionary and ontogenetic development of the carotid body is still understudied. Research aimed at studying the comparative morphology of the organ at different periods in the individual development of various animal species should play a crucial role in understanding the physiology of the carotid body. However, despite more than two centuries of study, the human carotid body remains poorly understood. There are many knowledge gaps in particular related to the antenatal development of this structure. The aim of our work is to study the morphological and immunohistochemical characteristics of the human carotid body in the antenatal and postnatal periods of development. We investigated the human carotid bodies from 1 embryo, 20 fetuses and 13 adults of different ages using samples obtained at autopsy. Immunohistochemistry revealed expression of βIII-tubulin and tyrosine hydroxylase in the type I cells and nerve fibers at all periods of ontogenesis; synaptophysin and PGP9.5 in the type I cells in some of the antenatal cases and all of the postnatal cases; 200 kDa neurofilaments in nerve fibers in some of the antenatal cases and all of the postnatal cases; and GFAP and S100 in the type II cells and Schwann cells in some of the antenatal cases and all of the postnatal cases. A high level of tyrosine hydroxylase in the type I cells was a distinctive feature of the antenatal carotid bodies. On the contrary, in the type I cells of adults, the expression of tyrosine hydroxylase was significantly lower. Our data suggest that the human carotid body may perform an endocrine function in the antenatal period, while in the postnatal period of development, it loses this function and becomes a chemosensory organ.

scholarly journals Gene expression and function of adenosine A2A receptor in the rat carotid body

2000 ◽  
Vol 279 (2) ◽  
pp. L273-L282 ◽  
Author(s):  
Shuichi Kobayashi ◽  
Laura Conforti ◽  
David E. Millhorn
Keyword(s):  
Carotid Body ◽  
Cellular Response ◽  
Cell Voltage ◽  
Type I ◽  
Type I Cells ◽  
Carotid Bodies ◽  
Intracellular Ca ◽  
Immunohistochemical Studies ◽  
I Cells ◽  
Receptor Mrnas

The present study was undertaken to determine whether rat carotid bodies express adenosine (Ado) A2A receptors and whether this receptor is involved in the cellular response to hypoxia. Our results demonstrate that rat carotid bodies express the A2A and A2B Ado receptor mRNAs but not the A1 or A3 receptor mRNAs as determined by reverse transcriptase-polymerase chain reaction. In situ hybridization confirmed the expression of the A2A receptor mRNA. Immunohistochemical studies further showed that the A2A receptor is expressed in the carotid body and that it is colocalized with tyrosine hydroxylase in type I cells. Whole cell voltage-clamp studies using isolated type I cells showed that Ado inhibited the voltage-dependent Ca2+ currents and that this inhibition was abolished by the selective A2A receptor antagonist ZM-241385. Ca2+ imaging studies using fura 2 revealed that exposure to severe hypoxia induced elevation of intracellular Ca2+ concentration ([Ca2+]i) in type I cells and that extracellularly applied Ado significantly attenuated the hypoxia-induced elevation of [Ca2+]i. Taken together, our findings indicate that A2A receptors are present in type I cells and that activation of A2Areceptors modulates Ca2+ accumulation during hypoxia. This mechanism may play a role in regulating intracellular Ca2+homeostasis and cellular excitability during hypoxia.

The identification of dopamine in the rabbit’s carotid body

1968 ◽  
Vol 170 (1019) ◽  
pp. 195-203 ◽  
Keyword(s):  
Carotid Body ◽  
Model System ◽  
Type I ◽  
Histochemical Technique ◽  
High Concentration ◽  
Metabolic Intermediate ◽  
Type I Cells ◽  
Carotid Bodies ◽  
Perivascular Nerves ◽  
I Cells

Application of the Falck & Hillarp histochemical technique to the rabbit carotid body reveals three fluorescent structures: brilliantly fluorescent Type I cells, varicose perivascular nerves, and weakly fluorescent non-varicose fibres. The Type I cell fluorescence is similar to that of a dopamine model system and has the appropriate activation and emission maxima. A catecholamine, identified as dopamine, has been extracted from homogenized carotid bodies, and estimated by the trihydroxyindole procedure. The concentration of the dopamine in the carotid body is estimated to be 20 to 40 μ g/g. This is very much greater than that of the noradrenaline present, of which there is about 1·5 μ g/g. The fluorescence of the Type I cells is attributed to the dopamine and it is suggested that the amine may be granule-bound. The unusually high concentration of dopamine could imply that it is not merely a metabolic intermediate in the carotid body.

scholarly journals Modulation of chronic hypoxia-induced chemoreceptor hypersensitivity by NADPH oxidase subunits in rat carotid body

2010 ◽  
Vol 108 (5) ◽  
pp. 1304-1310 ◽  
Author(s):  
L. He ◽  
X. Liu ◽  
J. Chen ◽  
B. Dinger ◽  
L. Stensaas ◽  
...  
Keyword(s):  
Nadph Oxidase ◽  
Carotid Body ◽  
Chronic Hypoxia ◽  
Body Tissue ◽  
Regulatory Subunit ◽  
Type I ◽  
Ros Production ◽  
Tissue Slices ◽  
Type I Cells ◽  
I Cells

Previous studies in our laboratory established that reactive oxygen species (ROS) generated by NADPH oxidase (NOX) facilitate the open state of a subset of K+ channels in oxygen-sensitive type I cells of the carotid body. Thus pharmacological inhibition of NOX or deletion of a NOX gene resulted in enhanced chemoreceptor sensitivity to hypoxia. The present study tests the hypothesis that chronic hypoxia (CH)-induced hypersensitivity of chemoreceptors is modulated by increased NOX activity and elevated levels of ROS. Measurements of dihydroethidium fluorescence in carotid body tissue slices showed that increased ROS production following CH (14 days, 380 Torr) was blocked by the specific NOX inhibitor 4-(2-amino-ethyl)benzenesulfonyl fluoride (AEBSF, 3 μM). Consistent with these findings, in normal carotid body AEBSF elicited a small increase in the chemoreceptor nerve discharge evoked by an acute hypoxic challenge, whereas after 9 days of CH the effect of the NOX inhibitor was some threefold larger ( P < 0.001). Evaluation of gene expression after 7 days of CH showed increases in the isoforms NOX2 (∼1.5-fold) and NOX4 (∼3.8-fold) and also increased presence of the regulatory subunit p47phox (∼4.2-fold). Involvement of p47phox was further implicated in studies of isolated type I cells that demonstrated an ∼8-fold and an ∼11-fold increase in mRNA after 1 and 3 days, respectively, of hypoxia in vivo. These findings were confirmed in immunocytochemical studies of carotid body tissue that showed a robust increase of p47phox in type I cells after 14 days of CH. Our findings suggest that increased ROS production by NOX enzymes in type I cells dampens CH-induced hypersensitivity in carotid body chemoreceptors.

Type I cell ROS kinetics under hypoxia in the intact mouse carotid body ex vivo: a FRET-based study

2015 ◽  
Vol 308 (1) ◽  
pp. C61-C67 ◽  
Author(s):  
A. Bernardini ◽  
U. Brockmeier ◽  
E. Metzen ◽  
U. Berchner-Pfannschmidt ◽  
E. Harde ◽  
...  
Keyword(s):  
Nadph Oxidase ◽  
Carotid Body ◽  
Ex Vivo ◽  
Specific Inhibitor ◽  
Type I ◽  
Oxygen Species ◽  
Nadph Oxidases ◽  
Intact Mouse ◽  
Type I Cells ◽  
I Cells

Reactive oxygen species (ROS) mainly originating from NADPH oxidases have been shown to be involved in the carotid body (CB) oxygen-sensing cascade. For measuring ROS kinetics, type I cells of the mouse CB in an ex vivo preparation were transfected with the ROS sensor construct FRET-HSP33. After 2 days of tissue culture, type I cells expressed FRET-HSP33 as shown by immunohistochemistry. In one population of CBs, 5 min of hypoxia induced a significant and reversible decrease of type I cell ROS levels ( n = 9 CBs; P < 0.015), which could be inhibited by 4-(2-aminoethyl)benzensulfonylfluorid (AEBSF), a highly specific inhibitor of the NADPH oxidase subunits p47phox and p67phox. In another population of CBs, however, 5 min of hypoxia induced a significant and reversible increase of ROS levels in type I cells ( n = 8 CBs; P < 0.05), which was slightly enhanced by administration of 3 mM AEBSF. These different ROS kinetics seemed to coincide with different mice breeding conditions. Type I cells of both populations showed a typical hypoxia-induced membrane potential (MP) depolarization, which could be inhibited by 3 mM AEBSF. ROS and MP closely followed the hypoxic decrease in CB tissue oxygen as measured with an O2-sensitive dye. We conclude that attenuated p47phox subunit activity of the NADPH oxidase under hypoxia is the physiological trigger for type I cell MP depolarization probably due to ROS decrease, whereas the observed ROS increase has no influence on type I cell MP kinetics under hypoxia.

Postnatal maturation of carotid body and type I cell chemoreception in the rat

1999 ◽  
Vol 276 (5) ◽  
pp. L875-L884 ◽  
Author(s):  
Owen S. Bamford ◽  
Laura M. Sterni ◽  
Michael J. Wasicko ◽  
Marshall H. Montrose ◽  
John L. Carroll
Keyword(s):  
Carotid Body ◽  
Carotid Sinus ◽  
Type I ◽  
Carotid Sinus Nerve ◽  
Postnatal Maturation ◽  
Cell Responses ◽  
Type I Cells ◽  
Intracellular Ca ◽  
I Cells

The site of postnatal maturation of carotid body chemoreception is unclear. To test the hypothesis that maturation occurs synchronously in type I cells and the whole carotid body, the development of changes in the intracellular Ca2+ concentration responses to hypoxia, CO2, and combined challenges was studied with fluorescence microscopy in type I cells and compared with the development of carotid sinus nerve (CSN) responses recorded in vitro from term fetal to 3-wk animals. Type I cell responses to all challenges increased between 1 and 8 days and then remained constant, with no multiplicative O2-CO2interaction at any age. The CSN response to hypoxia also matured by 8 days, but CSN responses to CO2 did not change significantly with age. Multiplicative O2-CO2interaction occurred in the CSN response at 2–3 wk but not in younger groups. We conclude that type I cell maturation underlies maturation of the CSN response to hypoxia. However, because development of responses to CO2 and combined hypoxia-CO2 challenges differed between type I cells and the CSN, responses to these stimuli must mature at other, unidentified sites within the developing carotid body.

Role of endothelin and endothelin A-type receptor in adaptation of the carotid body to chronic hypoxia

2002 ◽  
Vol 282 (6) ◽  
pp. L1314-L1323 ◽  
Author(s):  
J. Chen ◽  
L. He ◽  
B. Dinger ◽  
L. Stensaas ◽  
S. Fidone
Keyword(s):  
Carotid Body ◽  
Chronic Hypoxia ◽  
Acute Hypoxia ◽  
Critical Role ◽  
Immunocytochemical Staining ◽  
Type I ◽  
Type I Cells ◽  
I Cells

Chronic exposure in a low-Po 2 environment (i.e., chronic hypoxia, CH) elicits an elevated hypoxic ventilatory response and increased hypoxic chemosensitivity in arterial chemoreceptors in the carotid body. In the present study, we examine the hypothesis that changes in chemosensitivity are mediated by endothelin (ET), a 21-amino-acid peptide, and ETA receptors, both of which are normally expressed by O2-sensitive type I cells. Immunocytochemical staining showed incremental increases in ET and ETAexpression in type I cells after 3, 7, and 14 days of CH (380 Torr). Peptide and receptor upregulation was confirmed in quantitative RT-PCR assays conducted after 14 days of CH. In vitro recordings of carotid sinus nerve activity after in vivo exposure to CH for 1–16 days demonstrated a time-dependent increase in chemoreceptor activity evoked by acute hypoxia. In normal carotid body, the specific ETAantagonist BQ-123 (5 μM) inhibited 11% of the nerve discharge elicited by hypoxia, and after 3 days of CH the drug diminished the hypoxia-evoked discharge by 20% ( P < 0.01). This inhibitory effect progressed to 45% at day 9 of CH and to nearly 50% after 12, 14, and 16 days of CH. Furthermore, in the presence of BQ-123, the magnitude of the activity evoked by hypoxia did not differ in normal vs. CH preparations, indicating that the increased activity was the result of endogenous ET acting on an increasing number of ETA. Collectively, our data suggest that ET and ETA autoreceptors on O2-sensitive type I cells play a critical role in CH-induced increased chemosensitivity in the rat carotid body.

Interactions between hypoxia and hypercapnic acidosis on calcium signaling in carotid body type I cells

2000 ◽  
Vol 279 (1) ◽  
pp. L36-L42 ◽  
Author(s):  
Leonardo L. T. Dasso ◽  
Keith J. Buckler ◽  
Richard D. Vaughan-Jones
Keyword(s):  
Carotid Body ◽  
Neonatal Rat ◽  
Single Type ◽  
Type I ◽  
Hypercapnic Acidosis ◽  
Body Type ◽  
Type I Cells ◽  
Carotid Bodies ◽  
Intracellular Ca ◽  
I Cells

The effects of hypercapnic acidosis and hypoxia on intracellular Ca2+concentration ([Ca2+]i) were determined with Indo 1 in enzymatically isolated single type I cells from neonatal rat carotid bodies. Type I cells responded to graded hypoxic stimuli with graded [Ca2+]i rises. The percentage of cells responding was also dependent on the severity of the hypoxic stimulus. Raising CO2 from 5 to 10 or 20% elicited a significant increase in [Ca2+]i in the same cells as those that responded to hypoxia. Thus both stimuli can be sensed by each individual cell. When combinations of hypoxic and acidic stimuli were given simultaneously, the responses were invariably greater than the response to either stimulus given alone. Indeed, in most cases, the response to hypercapnia was slightly potentiated by hypoxia. These data provide the first evidence that the classic synergy between hypoxic and hypercapnic stimuli observed in the intact carotid body may, in part, be an inherent property of the type I cell.

Immunohistochemical demonstration of four subunits of neutrophil NAD(P)H oxidase in type I cells of carotid body

1995 ◽  
Vol 78 (5) ◽  
pp. 1904-1909 ◽  
Author(s):  
W. Kummer ◽  
H. Acker
Keyword(s):  
Hydrogen Peroxide ◽  
Carotid Body ◽  
Oxygen Sensor ◽  
Neutrophil Granulocytes ◽  
Type I ◽  
Body Type ◽  
Type I Cells ◽  
Carotid Bodies ◽  
Sensor Protein ◽  
I Cells

We demonstrate, by means of immunohistochemistry, that type I cells of human, guinea pig, and rat carotid bodies react with antisera raised against the subunits p22phox, gp91phox, p47phox, and p67phox of the NAD(P)H oxidase isolated from human neutrophil granulocytes. The findings support previous photometric studies that indicate that carotid body type I cells possess a putative oxygen sensor protein that is similar to the neutrophil NAD(P)H oxidase and consists of a hydrogen peroxide generating low-potential cytochrome b558 with cofactors regulating the electron transfer to oxygen.

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