Rat Carotid Body Glomus Cells Research Articles

Expression of p11 and TASK1 Channels in Rat Carotid Body Glomus Cells Subjected to Chronic Intermittent Hypoxia.

Chronic intermittent hypoxia (CIH) has been used as a model to mimic nocturnal apnea, which is associated with hypertension. One of the mechanisms for hypertension in patients with nocturnal apnea is an enhancement of the plasma membrane response to acute hypoxia in carotid body glomus cells. Hypoxia is known to induce depolarization via inhibiting TWIK-related acid-sensitive K+ (TASK) channels, one type of leak K+ channels, in glomus cells. The present experiment was undertaken to immunocytochemically investigate the effects of CIH on the expression and intracellular localization of TASK1 channels and p11 that critically affect the trafficking of TASK1 to the cell surface. The expression levels of TASK1 proteins and p11 and their intracellular localization in rat carotid body glomus cells were not noticeably affected by CIH, suggesting that the enhanced membrane response to acute hypoxia is not due to an increase in surface TASK channels.

Lack of influence of dexmedetomidine on rat glomus cell response to hypoxia, and on mouse acute hypoxic ventilatory response.

Background and Aims:There is a lack of basic science data on the effect of dexmedetomidine on the hypoxic chemosensory reflex with both depression and stimulation suggested. The primary aim of this study was to assess if dexmedetomidine inhibited the cellular response to hypoxia in rat carotid body glomus cells, the cells of the organs mediating acute hypoxic ventilatory response (AHVR). Additionally, we used a small sample of mice to assess if there was any large influence of subsedative doses of dexmedetomidine on AHVR.Material and Methods:In the primary study, glomus cells isolated from neonatal rats were used to study the effect of 0.1 nM (n = 9) and 1 nM (n = 13) dexmedetomidine on hypoxia-elicited intracellular calcium [Ca2%]i influx using ratiometric fluorimetry. Secondarily, whole animal unrestrained plethysmography was used to study AHVR in a total of 8 age-matched C57BL6 mice, divided on successive days into two groups of four mice randomly assigned to receive sub-sedative doses of 5, 50, or 500 μg.kg-1 dexmedetomidine versus control in a crossover study design (total n = 12 exposures to drug with n = 12 controls).Results:There was no effect of dexmedetomidine on the hypoxia-elicited increase in [Ca2%]i in glomus cells (a mean ± SEM increase of 95 ± 32 nM from baseline with control hypoxia, 124 ± 41 nM with 0.1 nM dexmedetomidine; P = 0.514). In intact mice, dexmedetomidine had no effect on baseline ventilation during air-breathing (4.01 ± 0.3 ml.g-1.min-1 in control and 2.99 ± 0.5 ml.g-1.min-1 with 500 μg.kg-1 dexmedetomidine, the highest dose; P = 0.081) or on AHVR (136 ± 19% increase from baseline in control, 152 ± 46% with 500 μg.kg-1 dexmedetomidine, the highest dose; P = 0.536).Conclusion:Dexmedetomidine had no effect on the cellular responses to hypoxia. We conclude that it unlikely acts via inhibition of oxygen sensing at the glomus cell. The respiratory chemoreflex effects of this drug remain an open question. In our small sample of intact mice, hypoxic chemoreflex responses and basal breathing were preserved.

Expression of p11 and Heteromeric TASK Channels in Rat Carotid Body Glomus Cells and Nerve Growth Factor-differentiated PC12 Cells.

TWIK-related acid-sensitive K+ (TASK) homomeric channels, TASK1 and TASK3, are present in PC12 cells. The channels do not heteromerize due plausibly to a lack of p11 protein. Single-channel recording reveals that most of the rat carotid body (CB) glomus cells express heteromeric TASK1-TASK3 channels, but the presence of p11 in glomus cells has not yet been verified. TASK1, but not TASK3, binds to p11, which has a retention signal for the endoplasmic reticulum. We hypothesized that p11 could facilitate heteromeric TASK1-TASK3 formation in glomus cells. We investigated this hypothesis in isolated immunocytochemically identified rat CB glomus cells. The findings were that glomus cells expressed p11-like immunoreactive (IR) material, and TASK1- and TASK3-like IR material mainly coincided in the cytoplasm. The proximity ligation assay showed that TASK1 and TASK3 heteromerized. In separate experiments, supporting evidence for the major role of p11 for channel heteromerization was provided in PC12 cells stimulated by nerve growth factor. p11 production took place there via multiple signaling pathways comprising mitogen-activated protein kinase and phospholipase C, and heteromeric TASK1-TASK3 channels were formed. We conclude that p11 is expressed and TASK1 and TASK3 heteromerize in rat CB glomus cells.

Hypoxia activates BK and Kv channels in rat carotid body glomus cells

Inhibition of BK and/or Kv has been proposed to be involved in hypoxia‐induced depolarization and subsequent elevation of intracellular [Ca2+] in carotid body glomus cells. However, depolarization and/or elevation of [Ca2+] are well known to activate BK and Kv, which would oppose glomus cell excitation. First we studied the effect of hypoxia on BK in cell‐attached patches, where the cell Em and intracellular [Ca2+] were allowed to change naturally in response to hypoxia. In cell‐attached patches (140 mM KCl in the pipette and 5 mM in the bath), hypoxia reversibly activated BK while causing inhibition of TASK. In cells partially depolarized by elevating extracellular [KCl] from 5 mM to 12–15 mM KCl, hypoxia also activated BK reversibly. In inside‐out patches showing different levels of BK activity, hypoxia produced no significant effect on BK. Kv could not be detected clearly in cell‐attached patches. Therefore, the role of Kv was tested indirectly by studying the effect of guangxitoxin (GxTX: Kv2 blocker) on hypoxia‐induced rise in intracellular [Ca2+]. GxTX inhibited the outward K+ current by ~50%, and enhanced the hypoxia‐induced elevation of intracellular [Ca2+] 2.3 fold, indicating that hypoxia activated Kv2. These results show that hypoxia activates BK and Kv in rat glomus cells, which helps to limit the hypoxia‐induced excitation of glomus cells.Support or Funding InformationSupported by NIH

Voltage- and receptor-mediated activation of a non-selective cation channel in rat carotid body glomus cells

A recent study showed that hypoxia activates a Ca2+-sensitive, Na+-permeable non-selective cation channel (NSC) in carotid body glomus cells. We studied the effects of mitochondrial inhibitors that increase Ca2+ influx via Ca2+ channel (Cav), and receptor agonists that release Ca2+ from endoplasmic reticulum (ER) on NSC. Mitochondrial inhibitors (NaCN, FCCP, H2S, NO) elevated [Ca2+]i and activated NSC. Angiotensin II and acetylcholine that elevate [Ca2+]i via the Gq-IP3 pathway activated NSC. However, endothelin-1 (Gq) and 5-HT (Gq) showed little or no effect on [Ca2+]i and did not activate NSC. Adenosine (Gs) caused a weak rise in [Ca2+]i but did not activate NSC. Dopamine (Gs) and γ-aminobytyric acid (Gi) were ineffective in raising [Ca2+]i and failed to activate NSC. Store-operated Ca2+ entry (SOCE) produced by depletion of Ca2+ stores with cyclopiazonic acid activated NSC. Our results show that Ca2+ entry via Cav, ER Ca2+ release and SOCE can activate NSC. Thus, NSC contributes to both voltage- and receptor-mediated excitation of glomus cells.

Role of BK in hypoxia‐induced elevation of [Ca2+]i in depolarized rat carotid body glomus cells

The controversial issue on the role of BK in hypoxia‐induced depolarization of glomus cells is not fully resolved. Some studies show that BK is closed in resting isolated cells, whereas others show that BK is open and inhibited by hypoxia. Here we tested the possibility that BK may be active in a subpopulation of glomus cells because they are partially depolarized and/or the basal [Ca2+]i is high and that hypoxia further depolarizes these cells by inhibiting BK. Patch‐clamp single channel recording was used to assess BK activity and fura‐2 fluorescence intensity was recorded to assess [Ca2+]i. Depolarizing glomus cells with 20 mM [KCl]o did not activate BK in cell‐attached patches, indicating that depolarization up to ~40 mV was not sufficient to activate BK in intact cells. In cells perfused with 20 mM [KCl]o, hypoxia (solution bubbled with 0% O2 gas) activated BK, presumably due to the additional depolarization caused by hypoxia. In cells bathed in 5 mM KClo, blocking BK with TEA (3 mM) or iberiotoxin (100 nM) did not increase the basal [Ca2+]i. In cells bathed in 15 and 20 mM KClo, TEA did not alter the averaged [Ca2+]i, although some variable responses were observed among cells. Hypoxia produced a large increase in [Ca2+]i in all cells. These results show that in both undepolarized and partially depolarized glomus cells, BK makes little or no contribution to hypoxia‐induced excitation of glomus cells, but BK may help to limit the depolarization produced by hypoxia in partially depolarized cells.

Activation of a Non‐Selective Cation Channel by Store‐Operated and Receptor‐Mediated Elevation of [Ca2+]i in Rat Carotid Body Glomus Cells

Our recent studies show that a non‐selective cation channel (NSC) is highly expressed in rat glomus cells. An increase in Ca2+ influx via the voltage‐dependent Ca2+ channel (VDCC) caused by depolarizing stimuli such as hypoxia, low pHo and high [KCl]o activated NSC. The effect of [Ca2+]i rise by mechanisms other than via VDCC on NSC activity is not yet known. Here we studied the role of store‐operated Ca2+ entry (SOCE) and receptor‐mediated [Ca2+]i rise on NSC activity. Patch‐clamp single channel recording and fura‐2 fluorescence signal were used. SOCE was produced by incubating cells in Ca2+‐free solution containing 1 uM thapsigargin and then adding Ca2+ back to the solution. The rapid rise in [Ca2+]i associated with SOCE caused a strong activation of NSC. Receptor‐mediated [Ca2+]i rise was produced using Angiotensin II (100 nM) and muscarine (10 uM), which stimulate the Gq‐PLC pathway and elevates [Ca2+]i in glomus cells. Both Ang II and muscarine produced a significant activation of NSC. Caffeine that causes release of Ca2+ from the endoplasimc reticulum also elevated [Ca2+]i and activated NSC. GABA, dopamine, endothelin‐1, 5‐HT did not elevate [Ca2+]i and did not activate NSC. Adenosine produced a weak elevation of [Ca2+]i but did not activate NSC. Our results indicate that a stimulus that produces an increase in [Ca2+]i, regardless of which signaling pathway (VDCC, SOCE and receptor‐mediated) is used, can activate NSC to help depolarize glomus cells.Support or Funding InformationSupported by NIH grant

Hydrogen sulfide and hypoxia-induced changes in TASK (K2P3/9) activity and intracellular Ca2+ concentration in rat carotid body glomus cells

Acute hypoxia depolarizes carotid body chemoreceptor (glomus) cells and elevates intracellular Ca2+ concentration ([Ca2+]i). Recent studies suggest that hydrogen sulfide (H2S) may serve as an oxygen sensor/signal in the carotid body during acute hypoxia. To further test such a role for H2S, we studied the effects of H2S on the activity of TASK channel and [Ca2+]i, which are considered important for mediating the glomus cell response to hypoxia. Like hypoxia, NaHS (a H2S donor) inhibited TASK activity and elevated [Ca2+]i. To inhibit the production of H2S, glomus cells were incubated (3h) with inhibitors of cystathionine-β-synthase and cystathionine-γ-lyase (dl-propargylglycine, aminooxyacetic acid, β-cyano-l-alanine; 0.3mM). SF7 fluorescence was used to assess the level of H2S production. The inhibitors blocked L-cysteine- and hypoxia-induced elevation of SF7 fluorescence intensity. In cells treated with the inhibitors, hypoxia produced an inhibition of TASK activity and a rise in [Ca2+]i, similar in magnitude to those observed in control cells. L-cysteine produced no effect on TASK activity or [Ca2+]i and did not affect hypoxia-induced inhibition of TASK and elevation of [Ca2+]i. These findings suggest that under normal conditions, H2S is not a major signal in hypoxia-induced modulation of TASK channels and [Ca2+]i in isolated glomus cells.

Hydrogen Sulfide and Hypoxia‐Induced Changes in K2P3/9 (TASK) current and [Ca2+]i in Rat Carotid Body Glomus Cells

Acute hypoxia depolarizes carotid body chemoreceptor (glomus) cells and elevates intracellular Ca2+ concentration ([Ca2+]i). Recent studies suggest that hydrogen sulfide (H2S) may serve as an oxygen sensor/signal in the carotid body during acute hypoxia. To further test such a role for H2S, we studied the effects of NaHS (a H2S donor) on the activity of TASK channel and intracellular [Ca2+]i, which are considered crucial for mediating the hypoxic response. Like hypoxia, NaHS inhibited TASK activity and elevated [Ca2+]i. To block the production of H2S, glomus cells were incubated (3 hr) with DL‐propargylglycine, aminooxyacetic acid, beta‐cyano‐L‐alanine (0.3 mM) that inhibit cystathionine‐beta‐synthase and cystathionine‐gamma‐lyase. Cell H2S production induced by L‐cysteine and hypoxia was strongly reduced by the inhibitors, as assessed using SF‐7 fluorescence. In cells treated with inhibitors, the hypoxia‐induced reduction of TASK activity and elevation of [Ca2+]i were similar in magnitude to those observed in untreated cells. In control or inhibitor‐treated cells, BK channel was not open at rest. These findings suggest that H2S does not mediate the hypoxia‐induced changes in TASK activity and [Ca2+]i in rat glomus cells.

Severe Hypoxia Activates BK channel in Rat Carotid Body Glomus Cells

Large-conductance Ca2+-activated K+ channel (BK) is highly expressed in rat carotid body glomus cells. Previous studies have reported that hypoxia inhibits BK, but the effect of hypoxia in intact glomus cells is not well defined. As hypoxia depolarizes glomus cells and elevates intracellular [Ca2+], we tested the hypothesis that hypoxia activates BK in intact cells. In cell-attached patches with 140 mM KCl in the pipette and 5 mM KCl in the bath solution, BK was always in the closed state, and hypoxia (0% O2) activated BK in ~10% of patches tested. In cell-attached patches, bath application of 1 mM NaCN also activated BK in some patches. When BK was first activated by depolarizing the patch membrane by changing the pipette potential or by perfusing the cells with high KCl (10-15 mM), hypoxia increased the BK activity reversibly. The stimulatory effect of hypoxia on BK was also observed in cell-attached patches with 5 mM KCl in the pipette, if the patch membrane was depolarized first. Study of the relationship among [Ca2+]i, cell Em and BK activity indicated that BK begins to activate when cell depolarizes ~30 mV and [Ca2+]i increases above ~500 nM (from a basal level of ~100 nM). Thus, severe hypoxia activates BK in glomus cells, and this presumably helps to limit over-excitation.

GAL-021 and GAL-160 are Efficacious in Rat Models of Obstructive and Central Sleep Apnea and Inhibit BKCa in Isolated Rat Carotid Body Glomus Cells.

GAL-021 and GAL-160 are alkylamino triazine analogues, which stimulate ventilation in rodents, non-human primates and (for GAL-021) in humans. To probe the site and mechanism of action of GAL-021 and GAL-160 we utilized spirometry in urethane anesthetized rats subjected to acute bilateral carotid sinus nerve transection (CSNTX) or sham surgery. In addition, using patch clamp electrophysiology we evaluated ionic currents in carotid body glomus cells isolated from neonatal rats. Acute CSNTX markedly attenuated and in some instances abolished the ventilatory stimulant effects of GAL-021 and GAL-160 (0.3 mg/kg IV), suggesting the carotid body is a/the major locus of action. Electrophysiology studies, in isolated Type I cells, established that GAL-021 (30 μM) and GAL-160 (30 μM) inhibited the BK(Ca) current without affecting the delayed rectifier K(+), leak K(+) or inward Ca(2+) currents. At a higher concentration of GAL-160 (100 μM), inhibition of the delayed rectifier K(+) current and leak K(+) current were observed. These data are consistent with the concept that GAL-021 and GAL-160 influence breathing control by acting as peripheral chemoreceptor modulators predominantly by inhibiting BK(Ca) mediated currents in glomus cells of the carotid body.

Possible Role of TRP Channels in Rat Glomus Cells.

Carotid body (CB) glomus cells depolarize in response to hypoxia, causing a [Ca(2+)](i) increase, at least in part, through activation of voltage-dependent channels. Recently, Turner et al. (2013) showed that mouse glomus cells with knockout of TASK1/3(-/-) channels have near-normal [Ca(2+)](i) response to hypoxia. Thus, we postulated that TRP channels may provide an alternate calcium influx pathway which may be blocked by the TRP channel antagonist, 2-APB (2-aminoethoxydiphenylborane). We confirmed that 2-APB inhibited the afferent nerve response to hypoxia, as previously reported (Lahiri S, Patel G, Baby S, Roy A (2009) 2-APB mediated effects on hypoxic calcium influx in rat carotid body glomus cells. FASEB 2009, Abstract, LB157; Kumar P, Pearson S, Gu Y (2006) A role for TRP channels in carotid body chemotransduction? FASEB J 20:A12-29). To examine the mechanism for this inhibition, we examined dissociated rat CB glomus cells for [Ca(2+)](i) responses to hypoxia, anoxia (with sodium dithionite), 20 mM K(+), NaSH, NaCN, and FCCP in absence/presence of 2-APB (100 μM). Also the effect of 2-APB on hypoxia and/or anoxia were investigated on NADH and mitochondria (MT) membrane potential. Our findings are as follows: (1) 2-APB significantly blocked the [Ca(2+)](i) increase in response to hypoxia and anoxia, but not the responses to 20 mM K(+). (2) The [Ca(2+)](i) responses NaSH, NaCN, and FCCP were significantly blocked by 2-APB. (3) Hypoxia-induced increases in NADH/NAD(+) and MT membrane depolarization were not effected by 2-APB. Thus TRP channels may provide an important pathway for calcium influx in glomus cells in response to hypoxia.

Effects of modulators of AMP-activated protein kinase on TASK-1/3 and intracellular Ca2+ concentration in rat carotid body glomus cells

Acute hypoxia depolarizes carotid body chemoreceptor (glomus) cells and elevates intracellular Ca2+ concentration ([Ca2+]i). Recent studies suggest that AMP-activated protein kinase (AMPK) mediates these effects of hypoxia by inhibiting the background K+ channels such as TASK. Here we studied the effects of modulators of AMPK on TASK activity in cell-attached patches. Activators of AMPK (1mM AICAR and 0.1–0.5mM A769662) did not inhibit TASK activity or cause depolarization during acute (10min) or prolonged (2–3h) exposure. Hypoxia inhibited TASK activity by ∼70% in cells pretreated with AICAR or A769662. Both AICAR and A769662 (15–40min) failed to increase [Ca2+]i in glomus cells. Compound C (40μM), an inhibitor of AMPK, showed no effect on hypoxia-induced inhibition of TASK. AICAR and A769662 phosphorylated AMPKα in PC12 cells, and Compound C blocked the phosphorylation. Our results suggest that AMPK does not affect TASK activity and is not involved in hypoxia-induced elevation of intracellular [Ca2+] in isolated rat carotid body glomus cells.

Perinatal hyperoxia exposure impairs hypoxia-induced depolarization in rat carotid body glomus cells

Chronic post-natal hyperoxia reduces the hypoxic ventilatory response by reducing the carotid body sensitivity to acute hypoxia as demonstrated by a reduced afferent nerve response, reduced calcium response of carotid body glomus cells and reduced catecholamine secretion in response to acute hypoxia. The present study examined whether hyperoxia alters the electrophysiological characteristics of glomus cells. Rats were treated with hyperoxia for 1 week starting at P1 or P7 and for 2 weeks starting at P1 followed by harvesting and dissociation of their carotid bodies for whole cell, perforated-patch recording. As compared to glomus cells from normoxia animals, hyperoxia treated cells showed a significant reduction in the magnitude of depolarization in response to hypoxia and anoxia, despite little change in the depolarizing response to 20mMK+. Resting cell membrane potential in glomus cells from rats exposed to hyperoxia from P1 to P15 and studied at P15 was slightly depolarized compared to other treatment groups and normoxia-treated cells, but conductance normalized to cell size was not different among groups. We conclude that postnatal hyperoxia impairs carotid chemoreceptor hypoxia transduction at a step between hypoxia sensing and membrane depolarization. This occurs without a major change in baseline electrophysiological characteristics, suggesting altered signaling or alterations in the relative abundance of different leak channel isoforms.

NAD(P)H autofluorescence induction by Compound C in rat carotid chemoreceptor cells

Compound C (CC) has been known as an inhibitor of AMP‐activated kinase (AMPK). Recently other roles of CC have been reported. CC inhibits hypoxia dependent HIF‐1α activation independent of AMPK but dependent on mitochondrial electron transport chain (ETC) (Emerling 07) and CC functions as an inhibitor of complex I of ETC (Chua 11). In addition, we have found that CC (4μM) produces ~10 fold increase in NAD(P)H autofluorescence (AutoFlu) as compared to hypoxia or cyanide in rat carotid body (CB) glomus cells. Autoflu was detected with 360/50 excitation(x) and 460/50 emission(m) filters. The effect of CC on the hypoxic [Ca2+]i response was observed by using 340&380x with 540/50m filter for fura‐2. Rat CB glomus cells were identified with rhodamine‐tagged peanut agglutinin.Results1) AutoFlu produced by CC is dose dependent, reaching a plateau at 0.04μM in both CB type cells. 2) The magnitude of AutoFlu induced by CC was greater in glomus cells compared to non‐glomus cells. 3) AutoFlu induced by 0.4μM CC was reduced ~50% by hypoxia but only < 10% by 20mM KCl. 4) CC did not increase calcium despite the increase in AutoFlu. 5) 0.4μM CC reduced the hypoxia‐induced [Ca2+]i increase by ~50%, but did not block 20mM KCl‐induced [Ca2+]i increase. These preliminary results suggest that CC induced significant NAD(P)H AutoFlu, which may significantly affect the measured [Ca2+]i response to hypoxia in glomus cells. (Funded by NIH HL54621)

Effects of modulators of AMPK on TASK K+ channel and [Ca2+] in rat carotid body glomus cells

Acute hypoxia depolarizes and elevates intracellular Ca2+ concentration ([Ca2+]i) in carotid body glomus cells. Recent studies suggest that AMP‐activated protein kinase (AMPK) mediates these effects of hypoxia by inhibiting background K+ channels such as TASK. To obtain additional evidence for the role of AMPK, we studied the effect of modulators of AMPK on TASK activity in cell‐attached patches of rat glomus cells. Perfusion of glomus cells for ~10 min with AICAR (1 mM) and A769662 (0.1 mM), two activators of AMPK, did not inhibit TASK activity or elicit depolarization. When cells were incubated with AICAR (1 mM), A769662 (0.1 mM), or DMSO (0.1%) for 2–3 hr, TASK activities in the three groups were also not significantly different. Furthermore, hypoxia inhibited TASK activity by 72% and 75% in cells pretreated with AICAR or A769662, respectively. Both AICAR and A769662 failed to increase [Ca2+]i in glomus cells. Compound C (40 μM) did not prevent hypoxia‐induced inhibition of TASK. AICAR and A769662 phosphorylated AMPKα in PC12 cells, and this was prevented by Compound C. Together, these findings suggest that inhibition of TASK by hypoxia is not mediated by AMPK. (supported by NIH and AHA)

Inhibition of rat carotid body glomus cells TASK-like channels by acute hypoxia is enhanced by chronic intermittent hypoxia

Chronic intermittent hypoxia (CIH), the main feature of obstructive sleep apnea, enhances carotid body (CB) chemosensory responses to acute hypoxia. In spite of that, the primary molecular target of CIH in the CB remains unknown. A key step of the hypoxic response in the CB is the chemoreceptor cell depolarization elicited by the inhibition of K+ channels. Thus, we tested the hypothesis that CIH potentiates the hypoxic-induced depolarization of rat CB chemoreceptor cells by enhancing the inhibition of a background K+ TASK-like channel. Membrane potential, single channel and macroscopic currents were recorded in the presence of TEA and 4-aminopyridine in CB chemoreceptor cells isolated from adult rats exposed to CIH. The CIH treatment did not modify the resting membrane properties but the hypoxic-evoked depolarization increased by 2-fold. In addition, the hypoxic inhibition of the TASK-like channel current was larger and faster in glomus cells from CIH-treated animals. This novel effect of CIH may contribute to explain the enhancing effect of CIH on CB oxygen chemoreception.

Characterization of an ATP-sensitive K+ channel in rat carotid body glomus cells

Carotid body glomus (CB) cells express different types of K(+) channels such as TASK, BK, and Kv channels, and hypoxia has been shown to inhibit these channels. Here we report the presence of a ∼72-pS channel that has not been described previously in CB cells. In cell-attached patches with 150 mM K(+) in the pipette and bath solutions, TASK-like channels were present (∼15 and ∼36-pS). After formation of inside-out patches, a 72-pS channel became transiently active in ∼18% of patches. The 72-pS channel was K(+)-selective, inhibited by 2-4 mM ATP and 10-100 μM glybenclamide. The 72-pS channel was observed in CB cells isolated from newborn, 2-3 week and 10-12 week-old rats. Reverse transcriptase-PCR and immunocytochemistry showed that Kir6.1, Kir6.2, SUR1 and SUR2 were expressed in CB glomus cells as well as in non-glomus cells. Acute hypoxia (∼15 mmHg O(2)) inhibited TASK-like channels but failed to activate the 72-pS channel in cell-attached CB cells. K(+) channel openers (diazoxide, pinacidil, levcromakalim), sodium cyanide and removal of extracellular glucose also did not activate the 72-pS channel in the cell-attached state. The hypoxia-induced elevation of intracellular [Ca(2+)] was unchanged by glybenclamide or diazoxide. NaCN-induced increase in [Ca(2+)] was not affected by 10 μM glybenclamide but inhibited by 100 μM glybenclamide. Acute glucose deprivation did not elevate [Ca(2+)] in the presence or absence of glybenclamide. These results show that an ATP-sensitive K(+) channel is expressed in the plasma membrane of CB cells, but is not activated by short-term metabolic inhibition. The functional relevance of the 72-pS channel remains to be determined.

Short-term Hypoxia Increases Tyrosine Hydroxylase Immunoreactivity in Rat Carotid Body

Neurochemical and morphological changes in the carotid body are induced by chronic hypoxia, leading to regulation of ventilation. In this study, we examined the time courses of changes in immunohistochemical intensity for tyrosine hydroxylase (TH) and cellular volume of glomus cells in rats exposed to hypoxia (10% O(2)) for up to 24 hr. Grayscale intensity for TH immunofluorescence was significantly increased in rats exposed to hypoxia for 12, 18, and 24 hr compared with control rats (p<0.05). The transectional area of glomus cells was not significantly different between experimental groups. The TH fluorescence intensity of the glomus cells exhibited a strong negative correlation with the transectional area in control rats (Spearman's rho = -0.70). This correlation coefficient decreased with exposure time, and it was lowest for the rats exposed to hypoxia for 18 hr (rho = -0.18). The histogram of TH fluorescence intensity showed a single peak in control rats. The peaks were gradually shifted to the right and became less pronounced in hypoxia-exposed rats, suggesting that a hypoxia-induced increase in TH immunoreactivity occurred uniformly in glomus cells. In conclusion, this study demonstrates that short-term hypoxia induces an increase in TH protein expression in rat carotid body glomus cells.

Hetero or homo, hypoxia has them all

Hetero or homo, hypoxia has them all