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The combined effects of hypoxia and hypercapnia on ventilation were demonstrated synergistic in mammalian. ( Neilson 1952). Although stimulus interaction between hypoxia and hypercapnia in peripheral chemoreceptors has been clearly demonstrated in experimental animals, there have been limited studies targeted in human. It is difficult to distinguish peripheral chemoreception from central chemoreception in steady-state ventilatory response to hypercapnia, since inspired hypercapnic gas stimulates both peripheral and central chemoreceptor. On the contrary, “two breaths method” has been used to estimate the peripheral chemoreceptor activity in human. In this method, transient alteration of ventilation with two breaths of hypoxic gas or hypercapnic gas may show the peripheral chemoreceptor activity. In the present study, we evaluate the ventilatory response to combined effects of hypoxia and hypercapnia in the peripheral chemosensitivity in healthy human by using two breaths method.

The respiratory response to extracellular acidosis by hypercapnia is mediated by central chemoreceptor neurons in the medulla oblongata [1]. There are actually two defined groups of respiratory neurons. The dorsal group of neurons is located in and near the nucleus of the tractus solitarius and their activity is regulated by changes in the arterial partial pressure of CO (Pco), O (Po) or H. The ventral group is a long column of neurons that extends through the nucleus ambiguous and retroambiguous in the ventrolateral medulla. In addition to reacting to peripheral stimuli, the ventral neurons detect changes in the H+ and/or CO concentrations in the cerebrospinal fluid (CSF) and brain interstitial fluid [2]. The capacity to detect these changes is called central chemosensitivity.

The carotid body is a primary sensory organ for arterial hypoxia. Chemosensory glomus cells in the carotid body release neurotransmitters, including ACh, in response to hypoxia. The release of neurotransmitters from the glomus cell, a putative chemoreceptor cell, appears to be triggered by an influx of calcium and subsequent increase in intracellular calcium ([Ca]i). Several reports indicate that L-type and some other types of voltage-gated calcium channels are responsible for neurotranmitter release from glomus cells (Gonzalez et al., 1994). These channels are activated by depolarization of the cell membrane. However, the speed and the degree of depolarization in glomus cells may not be sufficient to activate voltage-gated Ca channels at mild hypoxia (Chou et al., 1998), where afferent neural activity from the carotid body starts increasing. This discrepancy led us to search for other mechanisms which elevate [Ca]i followed by neurotransmitter release.

Previous investigators have suggested the existence of “CO sensors” in the lung and an important role of these receptors in detecting the increase in venous CO flux and in regulating ventilatory response to meet the metabolic demand during exercise (38). However, no direct and definitive evidence has been established in identifying the CO receptor in the lung.

In contrast to most other tissues, which exhibit considerable flexibility with respect to the nature of the substrates for their energy metabolism, the normal brain is restricted almost exclusively to glucose due to its distinguishing characteristics . Actual glucose utilization is 31 μmol/100 g tissue/min, in the normal, conscious human brain, indicating that glucose consumption is in excess for total oxygen consumption (Sokoloff, 1991). Although present in low concentration in brain (3.3 mmol/kg in rat), glycogen is a unique energy reserve for initiation of its metabolism. However, if glycogen concentration in the brain were the sole supply, normal energetic requirements would be maintained for less than 5 min (Sokoloff, 1991).

Airway sensory receptors regulate cardiopulmonary function by providing constant information about the mechanical and chemical status of the lung to the central nervous system (CNS). There are at least three airway sensor types: slowly adapting receptors (SARs), rapidly adapting receptors (RARs), and C-fiber receptors (CFRs). We recently identified additional A-delta fiber receptors in intact rabbits that are different from SARs and RARs. Having a high mechanical threshold, they respond to hypertonic saline and are termed high threshold A-delta receptors (HTARs). SARs and RARs monitor airway mechanical changes, whereas HTARs and CFRs sense chemical alterations and may serve as nociceptors. As with nociceptors in other tissue, the latter are activated during lung inflammatory processes. Also, the airway houses neuroendocrine cells aggregated in organoids called neuroepithelial bodies (NEBs). NEBs are richly innervated by nerve fibers from different origins. Similar in structure to the carotid bodies, NEBs are believed to be sensors, with at least some sensory fibers that have cell bodies in the nodose ganglia. Therefore, they may serve CNS reflex functions. Strategically located at airway bifurcations, NEBs may signal the chemical composition of or presence of irritants in the air. This study intends to explore the possibility that NEBs are associated with nociceptors.

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).

Pulmonary neuroepithelial bodies (NEB) are composed of innervated clusters of amine and peptide producing cells and are thought to function as hypoxia sensitive airway chemoreceptors. We have shown previously that the plasma membrane of rabbit fetal NEB in culture expresses an O sensing molecular complex composed of O sensitive K channel coupled to an O sensing protein (NADPH oxidase)(Nature, 1993;365:153). A Shaw-like, outward non-inactivating delayed- rectifier type K channel, was recorded from NEB cells in both culture and lung slices. This K channel was decreased by hypoxia (pO~20 mmHg), and was sensitive to TEA, 4-AP, and HO. Another whole cell K current recorded from NEB in culture exhibited electrophysiological characteristics of a slowly inactivating K current similar to the one described in Xenopus oocyte expressing Kv3.3a channel and this K current was increased by HO. Here we report findings on A-type K currents recorded from NEB in neonatal rabbit lung slice preparation. This slowly inactivating K current was inhibited by BDS-I (3 μM), specific blocker of Kv3.4 and rheteropodatoxin (HpTx-2; 0.2 μM), specific blocker of Kv4, and also sensitive to hypoxia. Using in situ hybridization method, mRNA for Kv3.4 and Kv4.3 was localized in NEB cells identified by immunostaining for serotonin. Expression of Kv3.4 and Kv4.3 proteins in NEB cells was confirmed by immunohistochemistry using specific antibodies. Multiple subtypes of voltage-dependent K current are expressed in NEB cells that may function as O-sensitive K channels.

Several neurotransmitters are present in the carotid body. These neurotransmitters are responsible to evoke action potentials in afferent nerve endings. They also modify the function of glomus cells, by binding to autoreceptors on glomus cells. We have previously demonstrated that ACh variably influences voltage-dependent K (Kv) current in cat glomus cells (Shirahata et al., 2002). Excitability of glomus cells are regulated by several types of K channels including voltage-dependent (Kv) channels (Buckler, 1999; Shirahata and Sham, 1999). Kv channels are activated with membrane depolarization, and play an essential role for repolarizing the cell membrane. Several studies have shown that the activity of Kv channels are modulated by neurotransmitters (Brown et al., 1997; Dong and White, 2003; Fukuda et al., 1988; Huang et al., 1993; Shi et al., 1999, 2004), and this type of modification may be important for fine tuning of the excitability of glomus cells. Recent studies have shown that the carotid body of DBA/2J inbred strain of mice demonstrates morphological similarities to cat glomus cells (Yamaguchi et al., 2003). Glomus cells of these mice responds to ACh (Yamaguchi et al., 2003) and express 4-aminopyridine sensitive and charybdotoxin sensitive components of Kv channels (Yamaguchi et al., 2004). In this study, we have investigated whether Kv channels in glomus cells of DBA/2J mice are also modified by ACh. Further, some underlying mechanisms of Kv current modulation by ACh was also investigated.

Carotid body (CB) chemoreceptor cells (CBCC) are involved in maintaining the homeostasis of O by detecting arterial blood PO and become activated when arterial PO decreases. In response to hypoxia, CBCC release neurotransmitters which excite the adjacent afferent nerve terminals of the carotid sinus nerve, increase their action potential and, via the central projections of the nerve to the brain stem, activate ventilation. The proposed cascade of transduction of hypoxia requires the presence of an oxygen sensor that is coupled to specific K channels. The decrease in the opening probability of these, leads to CBCC depolarization followed by Ca entry via voltagedependent Ca channels and consequently, the release of excitatory transmitters (5).However, the molecular identity of the O sensor and the mechanism coupling the oxygen sensor to the exocytotic machinery has thus far remained unclear.