Topical inhibition of nasal carbonic anhydrase affects the CO2 detection threshold in rats

Previous studies indicate that Long-Evans rats can be operantly trained to discriminate inspired CO(2) concentrations as low as 0.5%. This ability has been proposed to be due to the presence of CO(2)-sensitive olfactory receptors that contain the enzyme carbonic anhydrase (CA). The objectives of the present study were as follows: 1) to determine whether Zucker rats could be operantly conditioned to discriminate low concentrations of CO(2) from control air and 2) to determine the rats' CO(2) detection thresholds before and after nasal perfusion of mammalian Ringers or methazolamide, a CA inhibitor. Rats were operantly trained to discriminate between 25% CO(2) and control air (0% CO(2)) and were then subjected to various CO(2) concentrations (0.5-12.5%) to determine their CO(2) detection thresholds. The average (+/-standard error of mean) baseline CO(2) detection threshold of 7 Zucker rats was 0.48 +/- 0.07% CO(2), whereas the average CO(2) detection thresholds after nasal perfusion of either mammalian Ringers or 10(-2) M methazolamide were 1.41 +/- 0.30% and 5.92 +/- 0.70% CO(2), respectively. The average CO(2) detection threshold after methazolamide was significantly greater (P<0.0001) than the baseline detection threshold. These findings demonstrate that like Long-Evans rats, Zucker rats can be trained to discriminate low concentrations of CO(2) and that inhibition of nasal CA reduces the ability of the rats to detect low concentrations (3.5% and below) but not higher concentrations of CO(2) (12.5%). These results add to the growing evidence that olfactory neurons exhibiting CA activity are CO(2) chemoreceptors sensitive to physiological concentrations of CO(2).     

An assessment of nasal functions in control of breathing

Breathing pattern and steady-state CO2 ventilatory response during mouth breathing were compared with those during nose breathing in nine healthy adults. In addition, the effect of warming and humidification of the inspired air on the ventilatory response was observed during breathing through a mouthpiece. We found the following. 1) Dead space and airway resistance were significantly greater during nose than during mouth breathing. 2) The slope of CO2 ventilatory responses did not differ appreciably during the two types of breathing, but CO2 occlusion pressure response was significantly enhanced during nose breathing. 3) Inhalation of warm and humid air through a mouthpiece significantly depressed CO2 ventilation and occlusion pressure responses. These results fit our observation that end-tidal PCO2 was significantly higher during nose than during mouth breathing. It is suggested that a loss of nasal functions, such as during nasal obstruction, may result in lowering of CO2, fostering apneic spells during sleep.     

Alae nasi activation and nasal resistance in healthy subjects

To investigate the effect of alae nasi (AN) activation on nasal resistance, we monitored AN electromyographic (EMG) activity in 17 healthy subjects using surface electrodes placed on either side of the external nares and measured inspiratory nasal resistance utilizing the method of posterior rhinometry. With CO2 inhalation (6 subj), AN EMG activity increased as nasal resistance fell 23 +/- 5% (P less than 0.01). In the same subjects, voluntary flaring of the external nares also increased AN EMG and decreased nasal resistance by 29 +/- 5% (P less than 0.01). Nasal resistance was altered by nasal flaring and CO2 inhalation even after administration of a topical nasal vasoconstrictive spray (8 subj). In six subjects, voluntary nasal flaring or inhibition with the mouth closed produced a 21 +/- 12% change (P less than 0.01) in total airway resistance as measured by body plethysmography. We conclude that activation of the alae nasi will decrease nasal and total airway resistance during voluntary nasal flaring and during CO2 inhalation and thus should be considered in any studies of upper airway resistance.     

Effects of airway anesthesia on ventilatory responses to graded dead spaces and CO2

Ventilatory response to graded external dead space (0.5, 1.0, 2.0, and 2.5 liters) with hyperoxia and CO2 steady-state inhalation (3, 5, 7, and 8% CO2 in O2) was studied before and after 4% lidocaine aerosol inhalation in nine healthy males. The mean ventilatory response (delta VE/delta PETCO2, where VE is minute ventilation and PETCO2 is end-tidal PCO2) to graded dead space before airway anesthesia was 10.2 +/- 4.6 (SD) l.min-1.Torr-1, which was significantly greater than the steady-state CO2 response (1.4 +/- 0.6 l.min-1.Torr-1, P less than 0.001). Dead-space loading produced greater oscillation in airway PCO2 than did CO2 gas loading. After airway anesthesia, ventilatory response to graded dead space decreased significantly, to 2.1 +/- 0.6 l.min-1.Torr-1 (P less than 0.01) but was still greater than that to CO2. The response to CO2 did not significantly differ (1.3 +/- 0.5 l.min-1.Torr-1). Tidal volume, mean inspiratory flow, respiratory frequency, inspiratory time, and expiratory time during dead-space breathing were also depressed after airway anesthesia, particularly during large dead-space loading. On the other hand, during CO2 inhalation, these respiratory variables did not significantly differ before and after airway anesthesia. These results suggest that in conscious humans vagal airway receptors play a role in the ventilatory response to graded dead space and control of the breathing pattern during dead-space loading by detecting the oscillation in airway PCO2. These receptors do not appear to contribute to the ventilatory response to inhaled CO2.     

Encoding pheromonal signals in the mammalian vomeronasal system

The past few years have delivered substantial progress in understanding the molecular logic of the mammalian vomeronasal system. Selective expression of vomeronasal receptors and high response selectivity of vomeronasal receptor neurons suggest that pheromones are encoded by labeled lines at the level of the vomeronasal organ: each pheromonal compound is represented by the activation of a small and exclusive subset of receptor neurons. Labeled lines might be transferred to the accessory olfactory bulb through convergent connections. The key challenges ahead will be to identify the pheromonal ligands of the receptors and unravel the functional connectivity from the vomeronasal organ to the hypothalamus.     

The CO2/pH ventilatory drive in fish.

That ventilation in fish is driven by O2 has long been accepted. The O2 ventilatory drive reflects the much lower capacitance of water for O2 than for CO2, and is mediated by O2 receptors that are distributed throughout the gill arches and that monitor both internal and external O2 levels. In recent years, however, evidence has amassed in support of the existence of a ventilatory drive in fish that is keyed to CO2 and/or pH. While ventilatory responses to CO2/pH may be mediated in part by the O2 drive through CO2/pH-induced changes in blood O2 status, CO2/pH also appear to stimulate ventilation directly. The receptors involved in this pathway are as yet unknown, but the experimental evidence available to date supports the involvement of branchial CO2-sensitive chemoreceptors with an external orientation. Internally-oriented CO2-sensitive chemoreceptors may also be involved, although evidence on this point remains equivocal. In the present paper, the evidence for a CO2/pH-keyed ventilatory drive in fish will be reviewed.     

Rate of rise of intrapulmonary CO2 drives breathing frequency in garter snakes.

Garter snakes increase ventilation in response to elevated venous PCO2 without a concomitant rise in arterial PCO2 (Furilla et al. Respir. Physiol. 83: 47-60, 1991). Elevating venous PCO2 will increase the PCO2 gradient between pulmonary arterial blood and intrapulmonary gas during inspiration, leading to a greater rate of rise of intrapulmonary CO2 after inspiration. Because the lung contains CO2-sensitive receptors, I assessed the effect of the rate of rise of intrapulmonary CO2 on ventilation in unidirectionally ventilated snakes. CO2 concentration was altered using a digital gas mixer connected to a personal computer. Breathing frequency was highly correlated with the rate of rise intrapulmonary CO2 but only slightly affected by peak intrapulmonary CO2. On the other hand, tidal volume was more closely related to peak intrapulmonary CO2 than to the rate of rise of CO2. Bilateral pulmonary or cervical vagotomy nearly eliminated the ventilatory response associated with altered CO2 rise times but had little influence on the tidal volume response to the rate of rise of CO2. The mechanism whereby breathing frequency is controlled by the rate of rise of intrapulmonary CO2 is likely to originate with intrapulmonary chemoreceptors and may be important in the control of breathing during exercise.     

Maxillary palps are broad spectrum odorant detectors in Culex quinquefasciatus

A single type of olfactory sensilla on maxillary palps in many species of mosquitoes houses a very sensitive olfactory receptor neuron (ORN) for carbon dioxide reception. We performed extensive single sensillum recordings from this peg sensillum in Culex quinquefasciatus and have characterized the response threshold and kinetics for CO(2) reception, with a detection threshold less than the CO(2) concentration in the atmosphere. This ORN responded in a tonic mode to lower concentrations of CO(2), whereas higher concentrations generated a phasic-tonic mode of action potential firing. Sensillum potentials accurately represented the response magnitude and kinetics of carbon dioxide-elicited excitatory responses. Stimulation of these ORNs with human breath, a complex mixture of mosquito kairomones and up to 4.5% CO(2), elicited excitatory responses that were reliably detected by CO(2)-sensitive ORNs. Another ORN housed in these sensilla responded to 1-octen-3-ol and to various plant-derived compounds, particularly floral and green leaf volatiles. This ORN showed remarkable sensitivity to the natural enantiomer, (R)-(-)-1-octen-3-ol, rivaling pheromone-detecting ORNs in moths. Maximum neuronal response was elicited with a 10 ng dose. A biological, ecological role of maxillary palps in detection of plant- and nectar-related sources is proposed.     

Ventilatory response to exercise in aged runners breathing He-O2 or inspired CO2.

The ventilatory response to exercise below ventilatory threshold (VTh) increases with aging, whereas above VTh the ventilatory response declines only slightly. We wondered whether this same ventilatory response would be observed in older runners. We also wondered whether their ventilatory response to exercise while breathing He-O(2) or inspired CO(2) would be different. To investigate, we studied 12 seniors (63 +/- 4 yr; 10 men, 2 women) who exercised regularly (5 +/- 1 days/wk, 29 +/- 11 mi/wk, 16 +/- 6 yr). Each subject performed graded cycle ergometry to exhaustion on 3 separate days, breathing either room air, 3% inspired CO(2), or a heliox mixture (79% He and 21% O(2)). The ventilatory response to exercise below VTh was 0.35 +/- 0.06 l x min(-1) x W(-1) and above VTh was 0.66 +/- 0.10 l x min(-1) x W(-1). He-O(2) breathing increased (P < 0.05) the ventilatory response to exercise both below (0.40 +/- 0.12 l x min(-1) x W(-1)) and above VTh (0.81 +/- 0.10 l x min(-1) x W(-1)). Inspired CO(2) increased (P < 0.001) the ventilatory response to exercise only below VTh (0.44 +/- 0.10 l x min(-1) x W(-1)). The ventilatory responses to exercise with room air, He-O(2), and CO(2) breathing of these fit runners were similar to those observed earlier in older sedentary individuals. These data suggest that the ventilatory response to exercise of these senior runners is adequate to support their greater exercise capacity and that exercise training does not alter the ventilatory response to exercise with He-O(2) or inspired CO(2) breathing.     

Olfactory receptors and signalling elements in the Grueneberg ganglion

The Grueneberg ganglion (GG) is a cluster of neurones present in the vestibule of the anterior nasal cavity. Although its function is still elusive, recent studies have shown that cells of the GG transcribe the gene encoding the olfactory marker protein (OMP) and project their axons to glomeruli of the olfactory bulb, suggesting that they may have a chemosensory function. Chemosensory responsiveness of olfactory neurones in the main olfactory epithelium (MOE) and the vomeronasal organ (VNO) is based on the expression of either odorant receptors or vomeronasal putative pheromone receptors. To scrutinize its presumptive olfactory nature, the GG was assessed for receptor expression by extensive RT-PCR analyses, leading to the identification of a distinct vomeronasal receptor which was expressed in the majority of OMP-positive GG neurones. Along with this receptor, these cells expressed the G proteins Go and Gi, both of which are also present in sensory neurones of the vomeronasal organ. Odorant receptors were expressed by very few cells during prenatal and perinatal stages; a similar number of cells expressed adenylyl cyclase type III and G(olf/s), characteristic signalling elements of the main olfactory system. The findings of the study support the notion that the GG is in fact a subunit of the complex olfactory system, comprising cells with either a VNO-like or a MOE-like phenotype. Moreover, expression of a vomeronasal receptor indicates that the GG might serve to detect pheromones.     

Random expression of main and vomeronasal olfactory receptor genes in immature and mature olfactory epithelia of Fugu rubripes

Main olfactory receptor genes were isolated from a seawater fish, Fugu rubripes (pufferfish), and characterized. Two subfamilies of genes encoding seven transmembrane receptors were identified; one consists of five or more members, termed FOR1-1 to 5 of FOR1 subfamily, and the other appears to be a single copy gene, termed the FOR2 subfamily. FOR1 members show extremely high amino acid sequence similarities of about 95% to one another, and are distantly related to catfish-1 with the highest similarity of 37%. FOR2 shows 43% similarity to goldfish-A28. Phylogenically, both FOR members are categorized among pedigrees of the fish main olfactory receptor family outside the mammalian receptor family, although similarities between Fugu receptors and those of fresh-water fishes are lower than those among fresh-water fishes. In situ hybridization shows that both subfamilies of receptor genes are expressed randomly over the olfactory epithelium throughout all developmental stages, and no segregation of the signals was found. On the other hand, when three members of a vomeronasal olfactory receptor gene family, related to the Ca(2+)-sensing receptor, were used as probes, they were also randomly expressed over the same epithelium as the main olfactory receptors. This is in contrast to the expression profiles observed for zebrafish and goldfish, where the main or vomeronasal olfactory receptors are expressed in segregated patterns. It is thus suggested that the expression pattern of fish olfactory receptors varies depending on the species, although fish olfactory receptors are highly related to one another in their primary structures, and are phylogenically distinct from those of mammals.     

Ventilatory CO2 response in endurance-trained rats

A CO2 rebreathing technique was used to assess possible changes in the ventilatory response to CO2 in rats following a 14-week swim training program. Over the final 9 weeks, the rats swam 1 hr per day with a weight of 2.5% of the body weight attached to the tail. Ventilation was measured by a barometric method in awake, restrained rats in a total body plethysmography at CO2 concentrations of 0, 2, 4, 6, and 8%, with an initial O2 concentration of approximately 100%. Ventilation increased in the trained rats with increasing CO2 from 775ml . min-1 . kg-1 at 0% CO2 to 1,387 ml . min-1 . kg-1 at 8% CO2. This increase was a consequence of a 34% increase in tidal volume and a 32% increase in breathing frequency. In comparison with a group of sedentary control rats, there was a significantly higher ventilation and tidal volume at 0% CO2; however, this difference disappeared with increasing levels of CO2. A significantly lower resting heart rate was observed in the exercised (296 +/- 44 beats . min-1, mean +/- SD) compared to the sedentary control rats 380 +/- 42). It was concluded that, while the normal training response of resting bradycardia was observed following this duration and intensity of training, endurance swimming had no significant effect on the ventilatory response to CO2 in the rat.     

Pheromonal communication in amphibians

Pheromonal communication is widespread in salamanders and newts and may also be important in some frogs and toads. Several amphibian pheromones have been behaviorally, biochemically and molecularly identified. These pheromones are typically peptides or proteins. Study of pheromone evolution in plethodontid salamanders has revealed that courtship pheromones have been subject to continual evolutionary change, perhaps as a result of co-evolution between the pheromonal ligand and its receptor. Pheromones are detected by the vomeronasal organ and main olfactory epithelium. Chemosensory neurons express vomeronasal receptors or olfactory receptors. Frogs have relatively large numbers of vomeronasal receptors that are transcribed in both the vomeronasal organ and the main olfactory epithelium. Salamander vomeronasal receptors apparently are restricted to the vomeronasal organ. To date, no chemosensory ligands have been matched to vomeronasal receptors or olfactory receptors so it is unknown whether particular receptor types are (1) specialized for detection of pheromones versus other chemosignals, or (2) specialized for detection of volatile, nonvolatile, or water-borne chemosignals. Despite progress in understanding amphibian pheromonal communication, only a small fraction of amphibian species have been examined. Study of additional species of amphibians will indicate which traits related to pheromonal communication are evolutionarily conserved and which traits have diverged over time.     

Cold-induced changes in breathing pattern as a strategy to reduce respiratory heat loss

Thermoregulatory benefits of cold-induced changes in breathing pattern and mechanism(s) by which cold induces hypoventilation were investigated using male Holstein calves (1-3 mo old). Effects of ambient temperatures (Ta) between 4 and 18 degrees C on ventilatory parameters and respiratory heat loss (RHL) were determined in four calves. As Ta decreased, respiratory frequency decreased 29%, tidal volume increased 35%, total ventilation and RHL did not change, and the percentage of metabolic rate attributed to RHL decreased 26%. Total ventilation was stimulated by increasing inspired CO2 in six calves (Ta 4-6 degrees C), and a positive relationship existed between respiratory frequency and expired air temperature. Therefore, cold-exposed calves conserve respiratory heat by decreasing expired air temperature and dead space ventilation. Compared with thermoneutral exposure (16-18 degrees C), hypoventilation was induced by airway cold exposure (4-6 degrees C) alone and by exposing the body but not the airways to cold. Blocking nasal thermoreceptors with topical lidocaine during airway cold exposure prevented the ventilatory response but did not lower hypothalamic temperature. Hypothalamic cooling (Ta 16-18 degrees C) did not produce a ventilatory response. Thus, airway temperature but not hypothalamic temperature appears to control ventilation in cold-exposed calves.     

Chemical access to the vomeronasal organs of a plethodontid salamander

Plethodontid salamanders have unique nasolabial grooves that may function as “capillary tubes” to convey chemicals to the vomeronasal organ when these animals nose-tap. ³H-proline was placed at the base of these grooves in Plethodon cinereus, and autoradiography revealed large concentrations of radioactive material in the vomeronasal organs. There was no significant accumulation of radioactive material in the main olfactory epithelium. Salamanders with blocked nasolabial grooves lacked significant accumulation of material in their nasolabial grooves or vomeronasal epithelia, although some salamanders had radioactive material in the posterior portion of their vomeronasal organ that had entered through the internal nares. Anteriorly placed vomeronasal organs situated adjacent to the posterior limits of the nasolabial grooves may insure that nose-tapping primarily stimulates the vomeronasal sensory epithelium.     

Are geckos olfactory specialists?

Gekkonid lizards are shown to have well-developed nasal chemical senses. It is argued that they are unique among squamates so far studied in the degree of their olfactory (as opposed to vomeronasal) development. This contention is supported by evidence from the brain, nasal capsule, tongue, and experimental studies of behaviour. Limited evidence suggests that olfaction functions in food-finding and predator detection; vomerolfaction during investigation of novel stimuli and in reproduction. The conception of gekkonids as members of a 'visual Ascalabota' is not supported by these findings. Olfactory specialization makes geckos ideal subjects for tests of the Cowles and Phelan hypothesis of olfactory function and suggests that they might be better subjects than snakes for future studies of dual olfactory form, function and evolution in a nonmammalian lineage.     

Do lizards breathe through their mouths while running?

We measured the partitioning of airflow between nasal and oral circuits in five species of lizards before, during and after exercise. Expired gases were measured separately from the mouth and nose circuits in order to estimate the relative contribution of each circuit to ventilatory airflow. Nasal breathing dominates before exercise; however, during exercise the breathing pattern switched to oronasal expiration. Airflow averaged 30% oral expiration across all species during and after exercise. These results have important implications for the design of appropriate masks for respirometry in lizards. In order to ensure that all gases are captured, it is critically important that both the nose and mouth circuits are sampled.     

4种两栖爬行动物嗅器和犁鼻器的显微结构比较

用光镜观察了4种两栖爬行动物嗅器和犁鼻器的组织结构.结果显示,北方山溪鲵(Batrachuperus tibetanus)鼻囊内开始分化出犁鼻器,犁鼻器位于嗅器的腹外侧,但犁鼻器还不发达;隆肛蛙(Feirana quadranus)犁鼻器与嗅器虽然共同位于鼻囊内,但犁鼻器较为发达且其周围有发达的犁鼻腺,犁鼻器通过一细小管道与嗅器相通;秦岭蝮(Gloydius qinlingensis)和菜花烙铁头(Trimeresurus jerdonii)犁鼻腔与鼻腔已经完全分离形成两个独立的囊,而且鼻腔又进一步分化为嗅部与呼吸部.说明犁鼻器从有尾两栖动物开始出现,至无尾两栖类开始分化,到蛇类高度发达且成为一个独立器官.犁鼻器的形成是脊椎动物适应陆地生活的直接结果,是四足动物的特征之一.     

Stimulus parameters and temporal evolution of the olfactory evoked potential in rats

Evoked potentials were recorded from olfactory bulb, piriformcortex and scalp in urethane anesthetized rats in response tobrief odorant stimuli (amyl acetate, phenylethyl alcohol, eugenol)presented through a nasal cannula by means of a constant flowolfactometer. The effects of stimulus duration, nasal cannulaposition, flow rate, concentration and interstimulus intervalwere examined. The highest amplitude potentials were evokedby 10% amyl acetate at 20 ms duration, 1000 ml/min flow rateand a 60-s interstimulus interval with the stimulus deliveredat the nares. Odorant evoked potentials from deep within theolfactory bulb consisted of a triphasic wave with major componentsat 60 ms (P60), 90 ms (N90) and 140 ms (P140) with the lattertwo reversing polarity close to the surface of the bulb. Potentialsrecorded from layer I of piriform cortex were of similar amplitude,but opposite in polarity to the deep olfactory bulb potentials.Recordings from the skin over the nose elicited waveforms ofsimilar morphology to the deep olfactory bulb potentials, butone-quarter the amplitude and of opposite polarity The evokedpotentials changed with repetitive stimulation The N90 componentwas not present initially and only appeared after several stimuli.The appearance of the N90 component depended on the integrityof the olfactory peduncle. Thus, olfactory evoked potentialsto odorant stimuli reflect dynamic aspects of the encoding ofolfactory information dependent on connections between olfactorybulb and piriform cortex     

Olfactory and vomeronasal systems of caecilians (Amphibia: Gymnophiona)

The morphology of both the main nasal cavity and the vomeronasal organ differs among species representing six families of caecilians. The main nasal cavity is either divided or undivided. The vomeronasal organ differs in position (mediolateral, lateral), size (large vomeronasal organ in the aquatic species), and shape (mediolateral extension, vomeronasal organ with a lateral rostral projection). The great amount of respiratory epithelium of the main nasal cavity, the large vomeronasal organ, and its extensive innervation in typhlonectids may reflect both phylogeny and habitat adaptation, for these taxa are secondarily aquatic or semiaquatic and have several concomitant morphological and physiological modifications. The vomeronasal organ is associated with the caecilian tentacle as the tentacular ducts open into it. This association is further evidence for the involvement of the caecilian tentacle in vomeronasal chemoperception and may represent the mechanism by which these animals smell though the main nasal cavity is closed during burrowing or swimming. Labelings of primary olfactory and vomeronasal projections by means of horseradish peroxidase reaction reveal that the pattern of vomeronasal projections is similar in Ichthyophis kohtaoensis, Dermophis mexicanus, and Typhlonectes natans, even though T. natans possess stronger vomeronasal projections relative to olfactory projections than I. kohtaoensis and D. mexicanus. However, there are differences with respect to the patterns of olfactory projections. The olfactory projection of I. kohtaoensis is characterized by many displaced glomeruli. T. natans has the smallest olfactory projection. The nervus terminalis is associated with the olfactory system as shown by selective labelings of olfactory projections. Six characters potentially useful for phylogenetic analysis emerge from this study of comparative morphology. The characters were subjected to analysis using PAUP to see (1) if any resolution occurred and (2) if any groups were distinguished, whether they corresponded to phylogenetic arrangements based on other morphological characters. The characters are too few to produce nested dichotomous sets for all cases, but they do support the two typhlonectid genera examined and Dermophis and Gymnopis as sister taxa discrete from other groups, and they show that species within genera cluster together.