An increase in CO(2)/H(+) is a major stimulus for increased ventilation and is sensed by specialized brain stem neurons called central chemosensitive neurons. These neurons appear to be spread among numerous brain stem regions, and neurons from different regions have different levels of chemosensitivity. Early studies implicated changes of pH as playing a role in chemosensitive signaling, most likely by inhibiting a K(+) channel, depolarizing chemosensitive neurons, and thereby increasing their firing rate. Considerable progress has been made over the past decade in understanding the cellular mechanisms of chemosensitive signaling using reduced preparations. Recent evidence has pointed to an important role of changes of intracellular pH in the response of central chemosensitive neurons to increased CO(2)/H(+) levels. The signaling mechanisms for chemosensitivity may also involve changes of extracellular pH, intracellular Ca(2+), gap junctions, oxidative stress, glial cells, bicarbonate, CO(2), and neurotransmitters. The normal target for these signals is generally believed to be a K(+) channel, although it is likely that many K(+) channels as well as Ca(2+) channels are involved as targets of chemosensitive signals. The results of studies of cellular signaling in central chemosensitive neurons are compared with results in other CO(2)- and/or H(+)-sensitive cells, including peripheral chemoreceptors (carotid body glomus cells), invertebrate central chemoreceptors, avian intrapulmonary chemoreceptors, acid-sensitive taste receptor cells on the tongue, and pain-sensitive nociceptors. A multiple factors model is proposed for central chemosensitive neurons in which multiple signals that affect multiple ion channel targets result in the final neuronal response to changes in CO(2)/H(+).

Alterations in the partial pressure of carbon dioxide (P CO 2 ) and pH are known to change the activity of specialized central neurones referred to as chemosensitive neurones. We define a chemosensitive neurone as one whose firing rate is increased by an increase in CO 2 /H + . Such neurones have been localized to a number of brainstem centres involved in a variety of physiological functions including the regulation of cardiovascular and respiratory systems. These centres include the ventral medullary surface, the ventrolateral medulla (VLM), the solitary tract (NTS), the medullary raphe and the locus coeruleus (LC) (Nattie, 1995(Nattie, , 1998a(Nattie, ,b, 1999Richerson, 1995;Ritucci et al. 1998). The last of these regions, the LC, is located in the dorsal pons of the brainstem. It is the nucleus in the central nervous system with the highest number of noradrenergic neurones and has been associated with a number of physiological and behavioural functions including the sleep-wake cycle, feeding, cardiovascular and respiratory control, nociception, attention and learning (Hobson et al. 1975;Aston-Jones, 1985;Oyamada et al. 1998 Oyamada et al. 1998). The high degree of chemosensitivity makes this nucleus an ideal one for the study of the cellular pathways by which an increase in CO 2 /H + leads to an increase in neuronal discharge.Increased neuronal firing rate in response to high CO 2 /H + could be brought about by changes in three possible signals: extracellular pH (pH o ), intracellular pH (pH i ) or molecular CO 2 . There is evidence indicating that a change in pH i (intracellular acidification), rather than in pH o or molecular CO 2 , is the major signal responsible for the increase in firing rate seen in chemosensitive neurones (Lassen, 1990;Putnam, 2001;. Most cells respond to intracellular acidification with pH i recovery due to activation of pH-regulating membrane transporters. However, several studies have shown that neurones from various chemosensitive regions do not exhibit pH i recovery from intracellular acidification when pH o is also reduced (Ritucci et al. 1997;Putnam, 2001;), e.g. in response to hypercapnic acidosis (HA). For instance, neurones from the NTS and the VLM show sustained intracellular acidification without pH i recovery in response to hypercapnic acidosis _ , pH o 6.8) resulted in a slow intracellular acidification to a maximum fall of about 0.26 pH units and a 72 % increase in firing rate. For all treatments, the changes in pH i preceded or occurred simultaneously with the changes in firing rate and were considerably slower than the changes in pH o . In conclusion, an increased firing rate of LC neurones in response to acid challenges was best correlated with the magnitude and the rate of fall in pH i , indicating that a decrease in pH i is a major part of the intracellular signalling pathway that transduces an acid challenge into an increased firing rate in LC neurones.

III. Neuronal sensitivity to hyperoxia, hypercapnia, and inert gases at hyperbaric pressures. J Appl Physiol 95: 883-909, 2003; 10.1152/japplphysiol.00920.2002.-As ambient pressure increases, hydrostatic compression of the central nervous system, combined with increasing levels of inspired PO 2, PCO2, and N2 partial pressure, has deleterious effects on neuronal function, resulting in O 2 toxicity, CO2 toxicity, N2 narcosis, and high-pressure nervous syndrome. The cellular mechanisms responsible for each disorder have been difficult to study by using classic in vitro electrophysiological methods, due to the physical barrier imposed by the sealed pressure chamber and mechanical disturbances during tissue compression. Improved chamber designs and methods have made such experiments feasible in mammalian neurons, especially at ambient pressures Ͻ5 atmospheres absolute (ATA). Here we summarize these methods, the physiologically relevant test pressures, potential research applications, and results of previous research, focusing on the significance of electrophysiological studies at Ͻ5 ATA. Intracellular recordings and tissue PO 2 measurements in slices of rat brain demonstrate how to differentiate the neuronal effects of increased gas pressures from pressure per se. Examples also highlight the use of hyperoxia (Յ3 ATA O 2) as a model for studying the cellular mechanisms of oxidative stress in the mammalian central nervous system. anesthesia; carbon dioxide toxicity; free radicals; high-pressure nervous syndrome; membrane potential; nitrogen narcosis; oxidative stress; oxygen toxicity; polarographic oxygen electrode MOST HUMANS ARE PHYSIOLOGICALLY adapted to live and work near sea level, where ambient pressure is ϳ1 atmosphere absolute (ATA).1 Nonetheless, we exploit a continuum of barometric pressure (PB) ranging from the near vacuum of outer space, which we survive during Space Shuttle extravehicular activity by wearing a 4.3 lb./in.2 absolute (psia) pressure suit (30,300 ft. pressure equivalent, 0.29 ATA) (120, 220), down to the summit of Mt. Everest at 29,029 ft., where PB is 0.31 ATA (221), to ocean depths as great as 2,300 ft. of sea water (fsw), where PB increases to ϳ70 ATA (18). These situations are the pressure extremes of our inhabitable environment, which only relatively few highly trained individuals have ever occupied.Military personnel, medical personnel, and other humans, however, frequently encounter levels of hyperbaric pressure (i.e., Ͼ1 ATA) of lesser degrees in their normal work environments. Examples of moderate hyperbaric environments (e.g., Ͻ5 ATA) include the following: patients and medical attendants undergoing hyperbaric O 2 therapy (HBOT) (38,203); diving with an underwater breathing apparatus for recreational, professional (oil and salvage companies), and combat purposes (89); simulated dry and wet dives for hyperbaric research and dive training (217); and working in the compressed atmosphere of a subterranean environment (117). Abnormal work environments resulting from catastrophic accident...

. Hyperbaric oxygen and chemical oxidants stimulate CO2/H ϩ -sensitive neurons in rat brain stem slices. J Appl Physiol 95: 910-921, 2003. First published April 18, 2003 10.1152/japplphysiol.00864. 2002-Hyperoxia, a model of oxidative stress, can disrupt brain stem function, presumably by an increase in O2 free radicals. Breathing hyperbaric oxygen (HBO 2) initially causes hyperoxic hyperventilation, whereas extended exposure to HBO 2 disrupts cardiorespiratory control. Presently, it is unknown how hyperoxia affects brain stem neurons. We have tested the hypothesis that hyperoxia increases excitability of neurons of the solitary complex neurons, which is an important region for cardiorespiratory control and central CO2/H ϩ chemoreception. Intracellular recordings were made in rat medullary slices during exposure to 2-3 atm of HBO 2, HBO 2 plus antioxidant (Trolox C), and chemical oxidants (N-chlorosuccinimide, chloramine-T). HBO 2 increased input resistance and stimulated firing rate in 38% of neurons; both effects of HBO 2 were blocked by antioxidant and mimicked by chemical oxidants. Hypercapnia stimulated 32 of 60 (53%) neurons. Remarkably, these CO 2/H ϩ -chemosensitive neurons were preferentially sensitive to HBO2; 90% of neurons sensitive to HBO2 and/or chemical oxidants were also CO2/H ϩ chemosensitive. Conversely, only 19% of HBO2-insensitive neurons were CO2/H ϩ chemosensitive. We conclude that hyperoxia decreases membrane conductance and stimulates firing of putative central CO2/H ϩ -chemoreceptor neurons by an O2 free radical mechanism. These findings may explain why hyperoxia, paradoxically, stimulates ventilation. central chemoreception; reactive oxygen species; cardiorespiratory control; intracellular recording; hyperoxia THE CENTRAL NERVOUS SYSTEM (CNS) is especially sensitive to oxidative stress. For example, hyperoxia, which is a popular model of oxidative stress, induced by breathing high levels of oxygen at hyperbaric pressure [i.e., hyperbaric oxygen (HBO 2 )], can rapidly disrupt neural function and result in CNS O 2 toxicity (16). Neurological responses to hyperoxia vary, depending on the oxygen tension in the brain and the duration of exposure. For example, the CNS response to hyperoxia can range from moderate, but reversible, changes in neural activity (7,49), to violent and reversible seizures at higher levels of oxygen (16), to irreversible motor deficits and ultimately death at the highest dosages of hyperoxia (16). In each of these instances, the effects of hyperoxia on the CNS are thought to result from increased production and accumulation of O 2 free radicals and subsequent oxidation of cellular components vital to maintaining normal mechanisms of neuronal excitability (16).The cardiorespiratory centers of the brain stem, similarly, are sensitive to a broad range of inspired oxygen. Hypoxia increases alveolar ventilation, primarily by stimulation of the peripheral chemoreceptors (18), although a central stimulatory effect on ventilation has also been reported (47, 64). Paradoxica...

Hyperoxia is a popular model of oxidative stress. However, hyperoxic gas mixtures are routinely used for chemical denervation of peripheral O2 receptors in in vivo studies of respiratory control. The underlying assumption whenever using hyperoxia is that there are no direct effects of molecular O2 and reactive O2 species (ROS) on brain stem function. In addition, control superfusates used routinely for in vitro studies of neurons in brain slices are, in fact, hyperoxic. Again, the assumption is that there are no direct effects of O2 and ROS on neuronal activity. Research contradicts this assumption by demonstrating that O2 has central effects on the brain stem respiratory centers and several effects on neurons in respiratory control areas; these need to be considered whenever hyperoxia is used. This mini-review summarizes the long-recognized, but seldom acknowledged, paradox of respiratory control known as hyperoxic hyperventilation. Several proposed mechanisms are discussed, including the recent hypothesis that hyperoxic hyperventilation is initiated by increased production of ROS during hyperoxia, which directly stimulates central CO2 chemoreceptors in the solitary complex. Hyperoxic hyperventilation may provide clues into the fundamental role of redox signaling and ROS in central control of breathing; moreover, oxidative stress may play a role in respiratory control dysfunction. The practical implications of brain stem O2 and ROS sensitivity are also considered relative to the present uses of hyperoxia in respiratory control research in humans, animals, and brain stem tissues. Recommendations for future research are also proposed.