K+ channels expression in hypertension after arterial injury, and effect of selective Kv1.3 blockade with PAP-1 on intimal hyperplasia formation.

INTRODUCTION: K(+) channels are central to vascular pathophysiology. Previous results demonstrated that phenotypic modulation associates with a change in Kv1.3 to Kv1.5 expression, and that Kv1.3 blockade inhibits proliferation of VSMCs cultures. PURPOSE: To explore whether the Kv1.3 to Kv1.5 switch could be a marker of the increased risk of intimal hyperplasia in essential hypertension and whether systemic treatment with Kv1.3 blockers can prevent intimal hyperplasia after endoluminal lesion . METHODS: Morphometric and immunohistochemical analysis were performed in arterial segments following arterial injury and constant infusion of the Kv1.3 blocker PAP-1 during 28 days. Differential expression of K(+) channel genes was studied in VSMC from hypertensive (BPH) and normotensive (BPN) mice, both in control and after endoluminal lesion. Finally, the migration and proliferation rate of BPN and BPH VSMCs was explored in vitro. RESULTS: Changes in mRNA expression led to an increased Kv1.3/Kv1.5 ratio in BPH VSMC. Consistent with this, arterial injury in BPH mice induced a higher degree of luminal stenosis, (84 ± 4% vs. 70 ± 5% in BPN, p < 0.01), although no differences in migration and proliferation rate were observed in cultured VSMCs. The in vivo proliferative lesions were significantly decreased upon PAP-1 systemic infusion (18 ± 6% vs. 58 ± 20% with vehicle, p < 0.05). CONCLUSIONS: Hypertension leads to a higher degree of luminal stenosis in our arterial injury model, that correlates with a decreased expression of Kv1.5 channels. Kv1.3 blockers decreased in vitro VSMCs proliferation, migration, and in vivo intimal hyperplasia formation, pointing to Kv1.3 channels as promising therapeutical targets against restenosis.

DPPX modifies TEA sensitivity of the Kv4 channels in rabbit carotid body chemoreceptor cells.

Chemoreceptor cells from rabbit carotid body (CB) exhibit transient outward currents reversibly inhibited by low P(o2). Molecular and functional dissection of the components of these outward currents indicates that at least two different channels (Kv4.3 and Kv3.4) contribute to this current. Furthermore, several lines of evidence support the conclusion that Kv4 channel subfamily members (either Kv4.3 alone or Kv4.3/Kv4.1 heteromultimers) are the oxygen sensitive K channels (K(o2)) in rabbit CB chemoreceptor cells. However, the pharmacological characterization of these currents shows that they are almost completely blocked by high external TEA concentrations, while Kv4 channels have been shown to be TEA-insensitive. We hypothesized that the expression of regulatory subunits in chemoreceptor cells could modify TEA sensitivity of Kv4 channels. Here, we explore the presence and functional contribution of DPPX to K(o2) currents in rabbit CB chemoreceptor cells by using DPPX functional knockdown with siRNA. Our data suggest that DPPX proteins are integral components of K(o2) currents, and that their association with Kv4 subunits modulate the pharmacological profile of the heteromultimers.

Comparative gene expression profile of mouse carotid body and adrenal medulla under physiological hypoxia.

The carotid body (CB) is an arterial chemoreceptor, bearing specialized type I cells that respond to hypoxia by closing specific K+ channels and releasing neurotransmitters to activate sensory axons. Despite having detailed information on the electrical and neurochemical changes triggered by hypoxia in CB, the knowledge of the molecular components involved in the signalling cascade of the hypoxic response is fragmentary. This study analyses the mouse CB transcriptional changes in response to low PO2 by hybridization to oligonucleotide microarrays. The transcripts were obtained from whole CBs after mice were exposed to either normoxia (21% O2), or physiological hypoxia (10% O2) for 24 h. The CB transcriptional profiles obtained under these environmental conditions were subtracted from the profile of control non-chemoreceptor adrenal medulla extracted from the same animals. Given the common developmental origin of these two organs, they share many properties but differ specifically in their response to O2. Our analysis revealed 751 probe sets regulated specifically in CB under hypoxia (388 up-regulated and 363 down-regulated). These results were corroborated by assessing the transcriptional changes of selected genes under physiological hypoxia with quantitative RT-PCR. Our microarray experiments revealed a number of CB-expressed genes (e.g. TH, ferritin and triosephosphate isomerase) that were known to change their expression under hypoxia. However, we also found novel genes that consistently changed their expression under physiological hypoxia. Among them, a group of ion channels show specific regulation in CB: the potassium channels Kir6.1 and Kcnn4 are up-regulated, while the modulatory subunit Kcnab1 is down-regulated by low PO2 levels.

Ventilatory responses and carotid body function in adult rats perinatally exposed to hyperoxia.

Hypoxia increases the release of neurotransmitters from chemoreceptor cells of the carotid body (CB) and the activity in the carotid sinus nerve (CSN) sensory fibers, elevating ventilatory drive. According to previous reports, perinatal hyperoxia causes CSN hypotrophy and varied diminishment of CB function and the hypoxic ventilatory response. The present study aimed to characterize the presumptive hyperoxic damage. Hyperoxic rats were born and reared for 28 days in 55%-60% O2; subsequent growth (to 3.5-4.5 months) was in a normal atmosphere. Hyperoxic and control rats (born and reared in a normal atmosphere) responded with a similar increase in ventilatory frequency to hypoxia and hypercapnia. In comparison with the controls, hyperoxic CBs showed (1) half the size, but comparable percentage area positive to tyrosine hydroxylase (chemoreceptor cells) in histological sections; (2) a twofold increase in dopamine (DA) concentration, but a 50% reduction in DA synthesis rate; (3) a 75% reduction in hypoxia-evoked DA release, but normal high [K+]0-evoked release; (4) a 75% reduction in the number of hypoxia-sensitive CSN fibers (although responding units displayed a nearly normal hypoxic response); and (5) a smaller percentage of chemoreceptor cells that increased [Ca2+]1 in hypoxia, although responses were within the normal range. We conclude that perinatal hyperoxia causes atrophy of the CB-CSN complex, resulting in a smaller number of chemoreceptor cells and fibers. Additionally, hyperoxia damages O2-sensing, but not exocytotic, machinery in most surviving chemoreceptor cells. Although hyperoxic CBs contain substantially smaller numbers of chemoreceptor cells/sensory fibers responsive to hypoxia they appear sufficient to evoke normal increases in ventilatory frequency.

O(2) modulates large-conductance Ca(2+)-dependent K(+) channels of rat chemoreceptor cells by a membrane-restricted and CO-sensitive mechanism.

Hypoxic inhibition of large-conductance Ca(2+)-dependent K(+) channels (maxiK) of rat carotid body type I cells is a well-established fact. However, the molecular mechanisms of such inhibition and the role of these channels in the process of hypoxic transduction remain unclear. We have examined the mechanisms of interaction of O(2) with maxiK channels exploring the effect of hypoxia on maxiK currents recorded with the whole-cell and the inside-out configuration of the patch-clamp technique. Hypoxia inhibits channel activity both in whole-cell and in excised membrane patches. This effect is strongly voltage- and Ca(2+)-dependent, being maximal at low [Ca(2+)] and low membrane potential. The analysis of single-channel kinetics reveals a gating scheme comprising three open and five closed states. Hypoxia inhibits channel activity increasing the time the channel spends in the longest closed states, an effect that could be explained by a decrease in the Ca(2+) sensitivity of those closed states. Reducing maxiK channels with dithiothreitol (DTT) increases channel open probability, whereas oxidizing the channels with 2,2'-dithiopyridine (DTDP) has the opposite effect. These results suggest that hypoxic inhibition is not related with a reduction of channel thiol groups. However, CO, a competitive inhibitor of O(2) binding to hemoproteins, fully reverts hypoxic inhibition, both at the whole-cell and the single-channel level. We conclude that O(2) interaction with maxiK channels does not require cytoplasmic mediators. Such interaction could be mediated by a membrane hemoprotein that, as an O(2) sensor, would modulate channel activity.

Viral gene transfer of dominant-negative Kv4 construct suppresses an O2-sensitive K+ current in chemoreceptor cells.

Hypoxia initiates the neurosecretory response of the carotid body (CB) by inhibiting one or more potassium channels in the chemoreceptor cells. Oxygen-sensitive K(+) channels were first described in rabbit CB chemoreceptor cells, in which a transient outward K(+) current was reported to be reversibly inhibited by hypoxia. Although progress has been made to characterize this current with electrophysiological and pharmacological tools, no attempts have been made to identify which Kv channel proteins are expressed in rabbit CB chemoreceptor cells and to determine their contribution to the native O(2)-sensitive K(+) current. To probe the molecular identity of this current, we have used dominant-negative constructs to block the expression of functional Kv channels of the Shaker (Kv1.xDN) or the Shal (Kv4.xDN) subfamilies, because members of these two subfamilies contribute to the transient outward K(+) currents in other preparations. Delivery of the constructs into chemoreceptor cells has been achieved with adenoviruses that enabled ecdysone-inducible expression of the dominant-negative constructs and reporter genes in polycistronic vectors. In voltage-clamp experiments, we found that, whereas adenoviral infections of chemoreceptor cells with Kv1.xDN did not modify the O(2)-sensitive K(+) current, infections with Kv4.xDN suppressed the transient outward current in a time-dependent manner, significantly depolarized the cells, and abolished the depolarization induced by hypoxia. Our work demonstrate that genes of the Shal K(+) channels underlie the transient outward, O(2)-sensitive, K(+) current of rabbit CB chemoreceptor cells and that this current contributes to the cell depolarization in response to low pO(2).

Kvbeta1.2 subunit coexpression in HEK293 cells confers O2 sensitivity to kv4.2 but not to Shaker channels.

Voltage-gated K+ (KV) channels are protein complexes composed of ion-conducting integral membrane alpha subunits and cytoplasmic modulatory beta subunits. The differential expression and association of alpha and beta subunits seems to contribute significantly to the complexity and heterogeneity of KV channels in excitable cells, and their functional expression in heterologous systems provides a tool to study their regulation at a molecular level. Here, we have studied the effects of Kvbeta1.2 coexpression on the properties of Shaker and Kv4.2 KV channel alpha subunits, which encode rapidly inactivating A-type K+ currents, in transfected HEK293 cells. We found that Kvbeta1.2 functionally associates with these two alpha subunits, as well as with the endogenous KV channels of HEK293 cells, to modulate different properties of the heteromultimers. Kvbeta1.2 accelerates the rate of inactivation of the Shaker currents, as previously described, increases significantly the amplitude of the endogenous currents, and confers sensitivity to redox modulation and hypoxia to Kv4.2 channels. Upon association with Kvbeta1.2, Kv4.2 can be modified by DTT (1,4 dithiothreitol) and DTDP (2,2'-dithiodipyridine), which also modulate the low pO2 response of the Kv4.2+beta channels. However, the physiological reducing agent GSH (reduced glutathione) did not mimic the effects of DTT. Finally, hypoxic inhibition of Kv4.2+beta currents can be reverted by 70% in the presence of carbon monoxide and remains in cell-free patches, suggesting the presence of a hemoproteic O2 sensor in HEK293 cells and a membrane-delimited mechanism at the origin of hypoxic responses. We conclude that beta subunits can modulate different properties upon association with different KV channel subfamilies; of potential relevance to understanding the molecular basis of low pO2 sensitivity in native tissues is the here described acquisition of the ability of Kv4. 2+beta channels to respond to hypoxia.

Effects of almitrine bismesylate on the ionic currents of chemoreceptor cells from the carotid body.

Almitrine is a drug used in the treatment of hypoxemic chronic lung diseases such as bronchitis and emphysema because it is a potent stimulant of the carotid bodies in human and different animal species that produces a long-lasting enhancement of alveolar ventilation, ameliorating arterial blood gases. However, the mechanism of action of almitrine remains unknown. We investigated the effect of almitrine on ionic currents of chemoreceptor cells isolated from the carotid body of rat and rabbits by using the whole-cell and inside-out configurations of the patch-clamp technique. Almitrine at concentrations up to 10 microM did not affect whole-cell voltage-dependent K+, Ca2+, or Na+ currents in rat or rabbit cells. However, this concentration of almitrine significantly inhibited the Ca2+-dependent component of K+ currents in rat chemoreceptor cells. This effect of almitrine on the Ca2+-dependent component of K+ currents was investigated further at the single-channel level in excised patches in the inside-out configuration. In this preparation, almitrine inhibited the activity of a high-conductance (152 +/- 13 pS), Ca2+-dependent K+ channel by decreasing its open probability. The IC50 value of the effect was 0. 22 microM. The inhibitory effect of almitrine on Ca2+-dependent K+ channels also was observed in GH3 cells. We conclude that almitrine inhibits selectively the Ca2+-dependent K+ channel and that in rat chemoreceptor cells, this inhibition could represent an important mechanism of action underlying the therapeutic actions of the drug.

Mechanisms of sodium/calcium selectivity in sodium channels probed by cysteine mutagenesis and sulfhydryl modification.

A conserved lysine residue in the "P loop" of domain III renders sodium channels highly selective. Conversion of this residue to glutamate, to mimic the homologous position in calcium channels, enables Ca2+ to permeate sodium channels. Because the lysine-to-glutamate mutation converts a positively charged side chain to a negative one, it has been proposed that a positive charge at this position suffices for Na+ selectivity. We tested this idea by converting the critical lysine to cysteine (K1237C) in mu 1 rat skeletal sodium channels expressed in Xenopus oocytes. Selectivity of the mutant channels was then characterized before and after chemical modification to alter side-chain charge. Wild-type channels are highly selective for Na+ over Ca2+ (PCa/PNa < 0.01). The K1237C mutation significantly increases permeability to Ca2+ (PCa/PNa = 0.6) and Sr2+. Analogous mutations in domains I (D400C), II (E755C), and IV (A1529C) did not alter the selectivity for Na+ over Ca2+, nor did any of the domain IV mutations (G1530C, W1531C, and D1532C) that are known to affect monovalent selectivity. Interestingly, the increase in permeability to Ca2+ in K1237C cannot be reversed by simply restoring the positive charge to the side chain by using the sulfhydryl modifying reagent methanethiosulfonate ethylammonium. Single-channel studies confirmed that modified K1237C channels, which exhibit a reduced unitary conductance, remain permeable to Ca2+, with a PCa/PNa of 0.6. We conclude that the chemical identity of the residue at position 1237 is crucial for channel selectivity. Simply rendering the 1237 side chain positive does not suffice to restore selectivity to the channel.

Properties of ionic currents from isolated adult rat carotid body chemoreceptor cells: effect of hypoxia.

1. The electrical properties of chemoreceptor cells from neonatal rat and adult rabbit carotid bodies (CBs) are strikingly different. These differences have been suggested to be developmental and/or species related. To distinguish between the two possibilities, the whole-cell configuration of the patch-clamp technique was used to characterize the ionic currents present in isolated chemoreceptor cells from adult rat CBs. Since hypoxia-induced inhibition of O2-sensitive K+ currents is considered a crucial step in O2 chemoreception, the effect of hypoxia on the adult rat chemoreceptor cell currents was also studied. 2. Outward currents were carried mainly by K+, and two different components could be distinguished: a Ca(2+)-dependent K+ current (IK(Ca)) sensitive to Cd2+ and charybdotoxin (CTX), and a Ca(2+)-insensitive, voltage-dependent K+ current (IK(V)). IK(V) showed a slow voltage-dependent activation (time constant (tau) of 87.4 ms at -20 mV and 8.8 ms at +60 mV) and a very slow inactivation, described by the sum of two exponentials (tau 1 = 684 +/- 150 ms and tau 2 = 4.96 +/- 0.76 s at + 30 mV), that was almost voltage insensitive. The kinetic and pharmacological properties of IK(V) are typical of a delayed rectifier K+ channel. 3. Voltage-dependent Ca2+ currents (ICa) were present in nineteen of twenty-seven cells. TTX-sensitive Na+ currents were also observed in about 10% of the cells. 4. Low PO2 (< 10 mmHg) reduced the whole outward current amplitude by 22.17 +/- 1.96% (n = 27) at +20 mV. This effect was absent in the presence of Cd2+. Since low PO2 did not affect ICa, we conclude that hypoxia selectively blocks IK(Ca). 5. The properties of the currents recorded in adult rat chemoreceptor cells, including the specific inhibition of IK(Ca) by hypoxia, are similar to those reported in neonatal rat CB cells, implying that the differences between rat and rabbit chemoreceptor cells are species related.

Mechanisms of alpha2-adrenoceptor-mediated inhibition in rabbit carotid body.

We have used the in vitro preparation of the intact carotid body (CB) and isolated chemoreceptor cells to elucidate the distribution and function of alpha2-adrenoreceptors. The significance of the study lies in the fact that norepinephrine (NE), being the neurotransmitter of the sympathetic innervation to the CB, is also abundant in chemoreceptor cells. In intact CB whose catecholamine (CA) deposits had been labeled by prior incubation with the CA precursor [3H]tyrosine, the alpha2-antagonist yohimbine (10 microM) potentiated the low-PO2 (33 and 60 mmHg)-induced release of [3H]CA by 100 and 53%, respectively. Yohimbine (10 microM) and SKF-86466 (50 microM; another alpha2-antagonist) reversed the inhibition of the release of [3H]CA produced by the alpha2-receptor agonists clonidine and UK-14304 (10 microM). The increase in adenosine 3',5'-cyclic monophosphate produced by low PO2 was further augmented by yohimbine and nearly halved by UK-14304 and clonidine. In isolated chemoreceptor cells, UK-14304 and NE inhibited voltage-dependent Ca2+ currents by 28 and 32%, respectively. These results indicate that alpha2-receptors are present in chemoreceptor cells, where they reduce the release of [3H]CA. Inhibition of adenylate cyclase(s) and Ca2+ channels may be involved in this effect. Using intact CB from normal and chronically sympathectomized animals, we demonstrated a specific accumulation of [3H]NE in intraglomic sympathetic endings. Hypoxia (PO2 approximately 33 mmHg) did not elicit release of [3H]NE from the sympathetic endings, but high extracellular K+ (K+(e)) induced a release of [3H]NE that was inhibited by alpha2-agonists and augmented by alpha2-antagonists. These findings demonstrate that alpha2-receptors are also present in the sympathetic endings of the CB, where they modulate the release of NE. As a whole, this work provides a more detailed understanding of the role of the sympathetic innervation in the control of the CB chemoreceptor function, including the cellular mechanisms of the action of NE.

External pore residue mediates slow inactivation in mu 1 rat skeletal muscle sodium channels.

1. Upon depolarization, voltage-gated sodium channels assume non-conducting inactivated states which may be characterized as "fast' or "slow' depending on the length of the repolarization period needed for recovery. Skeletal muscle Na+ channel alpha-subunits expressed in Xenopus laevis oocytes display anomalous gating behaviour, with substantial slow inactivation after brief depolarizations. We exploited this kinetic behaviour to examine the structural basis for slow inactivation. 2. While fast inactivation in Na+ channels is mediated by cytoplasmic occlusion of the pore by III-IV linker residues, the structural features of slow inactivation are unknown. Since external pore-lining residues modulate C-type inactivation in potassium channels, we performed serial cysteine mutagenesis in the permeation loop (P-loop) of the rat skeletal muscle Na+ channel (mu 1) to determine whether similarly placed residues are involved in Na+ channel slow inactivation. 3. Wild-type and mutant alpha-subunits were heterologously expressed in Xenopus oocytes, and Na+ currents were recorded using a two-electrode voltage clamp. Slow inactivation after brief depolarizations was eliminated by the W402C mutation in domain I. Cysteine substitution of the homologous tryptophan residues in domains II, III and IV did not alter slow inactivation. 4. Analogous to the W402C mutation, coexpression of the wild-type alpha-subunit with rat brain Na+ channel beta 1-subunit attenuated slow inactivation. However, the W402C mutation imposed a delay on recovery from fast inactivation, while beta 1-subunit coexpression did not. We propose that the W402C mutation and the beta 1-subunit modulate gating through distinct mechanisms. 5. Removal of fast inactivation in wild-type alpha-subunits with the III-IV linker mutation I1303Q; F1304Q; M1305Q markedly slowed the development of slow inactivation. We propose that slow inactivation in Na+ channels involves conformational changes in the external pore. Mutations that affect fast and slow inactivation appear to interact despite their remote positions in the channel.

Depth asymmetries of the pore-lining segments of the Na+ channel revealed by cysteine mutagenesis.

We used serial cysteine mutagenesis to study the structure of the outer vestibule and selectivity region of the voltage-gated Na channel. The voltage dependence of Cd(2+) block enabled us to determine the locations within the electrical field of cysteine-substituted mutants in the P segments of all four domains. The fractional electrical distances of the substituted cysteines were compared with the differential sensitivity to modification by sulfhydryl-specific modifying reagents. These experiments indicate that the P segment of domain II is external, while the domain IV P segment is displaced internally, compared with the first and third domain P segments. Sulfhydryls with a steep voltage dependence for Cd(2+) block produced changes in monovalent cation selectivity; these included substitutions at the presumed selectivity filter, as well as residues in the domain IV P segment not previously recognized as determinants of selectivity. A new structural model is presented in which each of the P segments contribute unique loops that penetrate the membrane to varying depths to form the channel pore.

Enhancement of ionic current and charge movement by coexpression of calcium channel beta 1A subunit with alpha 1C subunit in a human embryonic kidney cell line.

1. Coexpression of the beta subunit with the alpha 1C subunit of the cardiac L-type Ca2+ channel has been shown to increase ionic current. To examine the mechanism of this increase, ionic and gating currents were measured in transiently transfected HEK293 cells. 2. Beta 1A subunit coexpression increased the maximal whole-cell conductance (Gmax) measured in 10 mM Ba2+ from 91 +/- 11 to 833 +/- 107 pS pF-1 without a change in the voltage dependence of activation (V1/2: -6.1 +/- 1.1 and -6.6 +/- 0.9 mV, respectively). 3. Gating currents were smaller in cells expressing only the alpha 1C subunit (only four out of eleven cells exhibited gating currents above the limits of detection, whereas eight out of eight beta 1A coexpressing cells had measurable gating currents). The gating currents were integrated to measure the intramembrane charge movement (Q). The ON charge movement (Qon) could be described by a Boltzmann distribution reaching a maximal value of Qon,max. 4. The mean ratio of Gmax: Qon,max increased from 99 +/- 6 to 243 +/- 30 pS fC-1 with beta 1A coexpression, demonstrating that the beta 1A subunit changes the gating of alpha 1C channels to favour the opening of the channels. However, this 2.5-fold change in the Gmax: Qon,max ratio explains less than half of the 9.2-fold increase in Gmax with beta 1A subunit coexpression. The major effect is due to a 3.7-fold increase in Qon,max, demonstrating that beta 1A subunit coexpression increases the number of functional surface membrane channels.

Control of ion flux and selectivity by negatively charged residues in the outer mouth of rat sodium channels.

1. The sodium channel has a ring of negatively charged amino acids on its external face. This common structural feature of cation-selective channels has been proposed to optimize conduction by electrostatic attraction of permeant cations into the channel mouth. We tested this idea by mutagenesis of mu1 rat skeletal sodium channels expressed in Xenopus oocytes. 2. Replacement of the external glutamate residue in domain II by cysteine reduces sodium current by decreasing single-channel conductance. While this effect can be reversed by the negatively charged sulfhydryl modifying reagent methanethiosulphonate ethylsulphonate (MTSES), the flux saturation behaviour cannot be rationalized simply by changes in the surface charge. 3. The analogous mutations in domains I, III and IV affect not only conductance but also selectivity. These changes in selectivity are only partially reversed by exposure to MTSES. 4. Our findings necessitate revision of prevailing concepts regarding the role of superficial negatively charged residues in the process of ion permeation. These residues do not act solely by electrostatic attraction of permeant ions, but instead may help to form ion-specific binding sites within the pore.

Structure of the sodium channel pore revealed by serial cysteine mutagenesis.

The pores of voltage-gated cation channels are formed by four intramembrane segments that impart selectivity and conductance. Remarkably little is known about the higher order structure of these critical pore-lining or P segments. Serial cysteine mutagenesis reveals a pattern of side-chain accessibility that contradicts currently favored structural models based on alpha-helices or beta-strands. Like the active sites of many enzymes of known structure, the sodium channel pore consists of irregular loop regions.

Cellular mechanisms of oxygen chemoreception in the carotid body.

The carotid bodies (CB) are arterial chemoreceptors that by sensing changes of arterial PO2, PCO2 and pH can initiate and modify ventilatory and cardiovascular reflexes in order to maintain PO2, PCO2 and pH within physiological levels. It is now generally accepted that the glomus or type I cells of the CB are the transducers of hypoxic stimuli, and relay chemosensory information to the brainstem via neurotransmitter release at synaptic contacts with afferent terminals of the carotid sinus nerve. This article reviews the mechanisms of the O2-sensing process at the cellular level. We consider first the transduction of the hypoxic stimulus, in which most of the experimental evidence currently favors a mechanism involving modulation of the electrical properties of type I cells. The last part of the article deals with the transmission of the stimulus between type I cells and afferent nerve terminals, and we present an overview on the issue of neurotransmission in the CB, summarizing the actions of the main neurotransmitters present in the organ.