CO2 chemosensitivity in Helix aspersa: three potassium currents mediate pH-sensitive neuronal spike timing

Elevated levels of carbon dioxide increase lung ventilation in Helix aspersa. The hypercapnic response originates from a discrete respiratory chemosensory region in the dorsal subesophageal ganglia that contains CO₂-sensitive neurons. We tested the hypothesis that pH-dependent inhibition of potassium channels in neurons in this region mediated the chemosensory response to CO₂. Cells isolated from the dorsal subesophageal ganglia retained CO₂ chemosensitivity and exhibited membrane depolarization and/or an increase in input resistance during an acid challenge. Isolated somata expressed two voltage-dependent potassium channels, an A-type and a delayed-rectifier-type channel (I_KA and I_KDR). Both conductances were inhibited during hypercapnia. The pattern of voltage dependence indicated that I_KA was affected by extracellular or intracellular pH, but the activity of I_KDR was modulated by extracellular pH only. Application of inhibitors of either channel mimicked many of the effects of acidification in isolated cells and neurons in situ. We also detected evidence of a pH-sensitive calcium-activated potassium channel (I_KCa) in neurons in situ. The results of these studies support the hypothesis that I_KA initiates the chemosensory response, and I_KDR and I_KCa prolong the period of activation of CO₂-sensitive neurons. Thus multiple potassium channels are inhibited by acidosis, and the combined effect of pH-dependent inhibition of these channels enhances neuronal excitability and mediates CO₂ chemosensory responses in H. aspersa. We did not find a single "chemosensory channel," and the chemosensitive channels that we did find were not unique in any way that we could detect. The protein "machinery" of CO₂ chemosensitivity is probably widespread among neurons, and the selection process whereby a neuron acts or does not act as a respiratory CO₂ chemosensor probably depends on the resting membrane potential and synaptic connectivity. carbon dioxide     

A computational analysis of central CO2 chemosensitivity in Helix aspersa

We created a single-compartment computer model of a CO₂ chemosensory neuron using differential equations adapted from the Hodgkin-Huxley model and measurements of currents in CO₂ chemosensory neurons from Helix aspersa. We incorporated into the model two inward currents, a sodium current and a calcium current, three outward potassium currents, an A-type current (I_KA), a delayed rectifier current (I_KDR), a calcium-activated potassium current (I_KCa), and a proton conductance found in invertebrate cells. All of the potassium channels were inhibited by reduced pH. We also included the pH regulatory process to mimic the effect of the sodium-hydrogen exchanger (NHE) described in these cells during hypercapnic stimulation. The model displayed chemosensory behavior (increased spike frequency during acid stimulation), and all three potassium channels participated in the chemosensory response and shaped the temporal characteristics of the response to acid stimulation. pH-dependent inhibition of I_KA initiated the response to CO₂, but hypercapnic inhibition of I_KDR and I_KCa affected the duration of the excitatory response to hypercapnia. The presence or absence of NHE activity altered the chemosensory response over time and demonstrated the inadvisability of effective intracellular pH (pH_i) regulation in cells designed to act as chemostats for acid-base regulation. The results of the model indicate that multiple channels contribute to CO₂ chemosensitivity, but the primary sensor is probably I_KA. pH_i may be a sufficient chemosensory stimulus, but it may not be a necessary stimulus: either pH_i or extracellular pH can be an effective stimuli if chemosensory neurons express appropriate pH-sensitive channels. The lack of pH_i regulation is a key feature determining the neuronal activity of chemosensory cells over time, and the balanced lack of pH_i regulation during hypercapnia probably depends on intracellular activation of pH_i regulation but extracellular inhibition of pH_i regulation. These general principles are applicable to all CO₂ chemosensory cells in vertebrate and invertebrate neurons. hypercapnia; potassium channels; computer modeling; central chemoreceptors     

Nucleus raphe obscurus modulates hypoglossal output of neonatal rat in vitro transverse brain stem slices

Nucleus raphéobscurus (NRo) modulates hypoglossal (XII) nerve motor output in the invitro transverse brain stem slice of neonatal rats (1-5 days old);chemical ablation of NRo and its focal CO₂ acidificationmodulated the bursting rhythm of XII nerves. We microinjected a 4.5 mMsolution of kainic acid into the NRo to disrupt cellular activity andobserved that XII nerve activity was temporarily abolished(n = 10). We also microinjected CO₂-acidified (pH = 6.00 ± 0.01) artificialcerebrospinal fluid (aCSF) into the NRo (n = 6), thepre-Bötzinger complex (PBC) (n = 6), as well as acontrol region in the lateral tegmental field equidistant to NRo, PBC,and the XII motor nuclei (n = 12). CO₂acidification of the control region had no effect on XII motor output.CO₂ acidification of the NRo significantly(P < 0.05) increased the burst discharge frequency ofXII nerves by 77%; integrated burst amplitude and burst durationincreased by 64% and 52%, respectively. CO₂ acidificationof the PBC significantly (P < 0.05) increased theburst discharge frequency of XII nerves by 65%, but neither integratedburst amplitude nor burst duration changed. These results demonstratethat chemical ablation of the NRo can abolish XII nerve bursting rhythmand that stimulation of the NRo with CO₂-acidified aCSF canexcite XII nerve bursting activity. From these observations, weconclude that, in transverse brain stem slices, the NRo containspH/CO₂-sensitive cells that modulate XII motor output.

     

Cellular mechanisms involved in CO(2) and acid signaling in chemosensitive neurons

An increase in CO₂/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 CO₂/H⁺ levels. The signaling mechanisms for chemosensitivity may also involve changes of extracellular pH, intracellular Ca²⁺, gap junctions, oxidative stress, glial cells, bicarbonate, CO₂, 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 Ca²⁺ 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 CO₂- 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 CO₂/H⁺. hypercapnia; brain stem; ventilation; peripheral chemoreceptor; glia; gap junction; glomus; channel; calcium; potassium; carbonic anhydrase; taste receptor; nociception     

Carbon Dioxide and Fruit Odor Transduction in Drosophila Olfactory Neurons. What Controls their Dynamic Properties?

We measured frequency response functions between odorants and action potentials in two types of neurons in Drosophila antennal basiconic sensilla. CO₂ was used to stimulate ab1C neurons, and the fruit odor ethyl butyrate was used to stimulate ab3A neurons. We also measured frequency response functions for light-induced action potential responses from transgenic flies expressing H134R-channelrhodopsin-2 (ChR2) in the ab1C and ab3A neurons. Frequency response functions for all stimulation methods were well-fitted by a band-pass filter function with two time constants that determined the lower and upper frequency limits of the response. Low frequency time constants were the same in each type of neuron, independent of stimulus method, but varied between neuron types. High frequency time constants were significantly slower with ethyl butyrate stimulation than light or CO₂ stimulation. In spite of these quantitative differences, there were strong similarities in the form and frequency ranges of all responses. Since light-activated ChR2 depolarizes neurons directly, rather than through a chemoreceptor mechanism, these data suggest that low frequency dynamic properties of Drosophila olfactory sensilla are dominated by neuron-specific ionic processes during action potential production. In contrast, high frequency dynamics are limited by processes associated with earlier steps in odor transduction, and CO₂ is detected more rapidly than fruit odor.     

Stomatal Opening in Light of Different Wavelengths: Effects of Blue Light Independent of Carbon Dioxide Concentration

Stomatal opening in Xanthium pennsylvanicum was found to besignificantly greater in blue light than in red. Experimentsin which leaves were placed in a closed system and allowed toestablish their own steady-state carbon dioxide concentrationshowed that when the CO₂ concentration was about the same asthat in red, opening was much greater in blue light. Blue lightof low intensity could cause as great an opening as red of higherintensity, even though the CO₂ concentration was much higherin blue. Stomatal opening in light is considered as involvingat least two reactions: (1) a response to the removal of CO₂by photosynthesis; (2) a response to blue light not dependenton the removal of CO₂. Blue light became increasingly effective, relative to red, asthe length of night was increased over the range 2 to 14 hours.This might, in part, explain previously observed effects ofnight length on rate of opening in light. The initial very rapid phase of closure in darkness appearedto be independent of CO₂ accumulation, for it was not preventedby flushing the intercellular spaces with air free of CO₂. Itis suggested that closure in darkness, like opening in light,should be considered as involving components both dependentupon, and independent of, CO₂ concentration.     

Effects of Elevated CO2 and Simulated Seasonal Changes in Temperature on the Species Composition and Growth Rates of Pasture Turves

Large turves from a ryegrass/white clover based pasture wereexposed to 350 or 700 µl l^-1 CO₂ for a period of 217 din controlled environment rooms. The temperature was increasedduring the experiment from 10/4 °C day/night to 16/10 °Cand finally to 22/16 °C. The turves were cut to a heightof 2 cm at intervals and growth rates calculated from the regrowth. Growth rates over the duration of the experiment were 8% higherat elevated CO₂; the difference between CO₂ treatments beingstatistically significant only at the highest temperature. Speciescomposition of the turves at 350 µl l^-1 CO₂ showed seasonalchanges similar to those measured in the field. The effect ofCO₂ was to exaggerate the normal decline of ryegrass at warmertemperatures and increase the proportion of white clover. About30% of the total growth rate was from other species (notablyBromus hordeaceus L. and Poa trivialis L.) and this fractionwas similar between CO₂ levels. Root mass was measured at theend of the experiment and was 50% higher at elevated CO₂. The modest above-ground response to CO₂ was a result of CO₂stimulation occurring only at the higher temperature. Becauseof the CO₂ x temperature interaction, the effect of CO₂ in temperateregions will be seasonal. When this is matched with seasonalgrowth patterns of herbage species, a complex response of pasturecommunities to CO₂ is possible. In our case, white clover wasgrowing most strongly during the period of greatest CO₂ stimulationand consequently its growth was enhanced more than that of ryegrass;however, the cooler season growth of ryegrass gives it a temporalniche which is little affected by CO₂ and this may be importantfor ryegrass stability if it is an inherently poor responderto CO₂. The results indicate that for temperate species theeffects of competition at elevated CO₂ cannot be easily determinedfrom experiments conducted at a single temperature.Copyright1994, 1999 Academic Press CO₂ enrichment, seasonal growth, species composition, turves, Trifolium repens L., Lolium perenne L., climate change     

Stomatal responses to light and CO2 in greening wheat leaves

The development of two types of stomatal transpiration, oneinduced by light (light-induced stomatal transpiration) andthe other induced by CO₂-free air in the dark (CO₂-sensitivestomatal transpiration), in greening leaves of wheat (Triticumaestivum L.) was studied in respect to the development of CO₂uptake and chlorophyll formation. Light-induced stomatal transpirationwas not observed at all in etiolated leaves and was generatedafter 3 hr of illumination for greening, when the activity ofCO₂ uptake was generated. CO₂-sensitive stomatal transpirationwas low in etiolated leaves and started to increase at the sametime during greening as the start of CO₂ uptake. The activitiesof both light-induced and CO₂-sensitive stomatal transpirationincreased as the activity of CO₂ uptake and the chlorophyllcontent increased. Pre-illumination of etiolated leaves for1 min followed by 4 hr of dark incubation eliminated the lagfor the development of the two types of stomatal transpirationand CO₂ uptake. (Received September 4, 1978; )     

Studies on the Mechanisms of Action of the Antitranspirant Phenylmercuric Acetate, and its Penetration into the Mesophyll

Experiments have examined the effect of phenylmercuric acetate(PMA) on the guard cells of Commelina communis. In one series,PMA was supplied to the leaf surface; after different time intervalsthe epidermis was removed and the ability of the stomata toopen was determined. In the other series, different concentrationsof PMA were included in the medium used for inccubating epidermalstrips with which ion-stimulated stomatal opening was assayed.At concentrations of 10^-54 M and above the effect of PMA wassevere and the structural integrity of the guard cells was affected;they were unable to accumulate neutral red. At concentrationsarpound 10^-6 M the guard cells were less affected and PMA broughtabout a transient stimulation of stomatal opening by releasingsubsidiary-cell turgor pressure. A solution of 5 x 10^-4 M PMA applied to leaves reduced by halfthe photosynthetic ¹⁴CO₂ incorporation into C. communis mesophyll.In Zea mays it increased the CO₂ compensation point and alsothe resistance to diffusion in the gas phase (R_A, but therewas a proportionately greater increase in the apparent liquidphase resistance (R_t). This direct inhibition of mesophyll photosynthesisundermines one of the major objectives of applying anatitranspirants,and for this reason it is suggested that PMA is unsuitable forgeneral application to crops.     

Effects of Elevated Carbon Dioxide on Plant Biomass Production and Competition in a Simulated Neutral Grassland Community

Using open-top chambers, four prominent species (Lolium perenne,Cynosurus cristatus, Holcus lanatusandAgrostis capillaris) ofIrish neutral grasslands were grown at ambient and elevated(700 µmol mol^-1) atmospheric CO₂for a period of 8 months.The effects of interspecific competition on plant responsesto CO₂enrichment were investigated by growing the species ina four-species mixture. The results indicate that the speciesdiffer in their ability to respond to elevated CO₂. CO₂-enrichmenthad the largest effect on the biomass production ofH. lanatus,but substantial stimulations in biomass production were alsofound for the other three species. The CO₂-stimulation of biomassproduction forH. lanatuswas accompanied by increased tillering.In addition, reductions in specific leaf area were found forall species. Exposure to elevated CO₂increased the communitybiomass of the four-species mixture. This increase can be mainlyattributed to a significant increase in the biomass ofH. lanatusatelevated CO₂. No statistically-significant changes in speciescomposition of community biomass were found. However,H. lanatusdidincrease its share of community biomass at each of the harvests,with the other three species, mainlyL. perenne, suffering lossesin their shares at elevated CO₂. The results show that: (1)the species varied in their response to elevated CO₂; and (2)species composition in natural plant communities is likely tochange at elevated CO₂, but these changes may occur rather slowly.Much longer periods of exposure to elevated atmospheric CO₂maybe required to permit detection of significant changes in speciescomposition.Copyright 1998 Annals of Botany Company Carbon dioxide (CO₂) enrichment, competition, Lolium perenne,Cynosurus cristatus, Holcus lanatus, Agrostis capillaris, biomass, specific leaf area, tillering.     

Changes in the Morphology of Roots and Leaves of Carob Seedlings Induced by Nitrogen Source and Atmospheric Carbon Dioxide

Carob seedlings were grown hydroponically for 9 weeks under360 and 800 µl l^-1CO₂. One of two nitrogen sources, nitrateor ammonium, was added to the nutrient medium at concentrationsof 3 mol m^-3. Root systems of the developing plants suppliedwith nitrate compared to those supplied with ammonium were characterizedby:(a)more biomass on the lower part of the root;(b)fewer lateralroots of first and second order;(c)longer roots;(d)higher specificroot length;(e)a smaller root diameter. The morphology of theroot systems of nitrate-fed plants changed in the presence ofelevated carbon dioxide concentrations, resembling, more closely,that of ammonium-fed plants. Total leaf area was higher in ammonium-than in nitrate-fed plants. Nitrate-fed plants had greater totalleaf area in the presence of high carbon dioxide than in normalCO₂, due to an increase in epidermal cell size that led to developmentof larger leaflets with lower stomatal frequency. The observedchanges in the morphology of roots and shoots agreed with theresults observed for total biomass production. Nitrate-fed plantsincreased their biomass production by 100% in the presence ofelevated CO₂compared to 15% in ammonium-fed plants, indicatingthat the response of carob to high CO₂concentrations is verydependent on the nitrogen source. Under elevated CO₂, nitrate-grownplants had a larger content of sucrose in both roots and shoots,while no significant difference was observed in the contentof sucrose in ammonium-grown plants, whether in ambient or enrichedcarbon dioxide. Hence, the differences in soluble carbohydratecontents can, at least partly, account for differences in rootand shoot morphology.Copyright 1997 Annals of Botany Company Ceratonia siliquaL.; carob; ammonium; carbohydrate; carbon dioxide; nitrate; morphology; sucrose     

Photosynthesis and Diffusion Conductance of the Valencia Orange Fruit under Field Conditions

CO₂ uptake and diffusion conductance of Valencia orange fruits(Citrus sinensis L. Osbeck) were measured in the field duringthe growing season of 1977/78 to ascertain if, as in the leaf,stomata control photosynthesis and transpiration under changingenvironmental conditions. Measurements were made on 15 yearold trees grown in a sandy loam soil and receiving either adry or a wet treatment. Fruit diffusive conductance was measuredwith a modified water vapour diffusion conductance meter andgross photosynthesis was measured with a ¹⁴CO₂ uptake meter.Photosynthetically active radiation (PAR) was measured witha quantum sensor. Fruits exposed to light assimilated CO₂ ata rate which was 25–50% of that assimilated by leaves.The uptake was dependent on fruit size, PAR, chlorophyll content,and on diffusive conductance of the fruit epidermis. Epidermalconductance showed a diurnal trend which was similar in shapeto that of the leaf except in the late afternoon. Cuticularconductance of the fruit was calculated and ranged between 0.22and 0.30 mm s^–1. It was speculated that the CO₂ uptakeby the fruit could support the growth of flavedo cell layerswhen exposed to light. Dry soil caused an increase in the ¹⁴CO₂uptake by fruit possibly caused by the increased potential areaof the stomatal opening per unit of fruit surface area.     

Compensatory Roles of Nitrogen Uptake and Photosynthetic N-use Efficiency in Determining Plant Growth Response to Elevated CO2: Evaluation Using a Functional Balance Model

We used a modified functional balance (FB) model to predictgrowth response of Helianthus annuus L. to elevated CO₂. Modelpredictions were evaluated against measurements obtained twiceduring the experiment. There was a good agreement between modelpredictions of relative growth rate (RGR) responses to elevatedCO₂and observations, particularly at the second harvest. Themodel was then used to compare the relative effects of biomassallocation to roots, nitrogen (N) uptake and photosyntheticN-use efficiency (PNUE) in determining plant growth responseto elevated CO₂. The model predicted that a rather substantialincrease in biomass allocation to root growth had little effecton whole plant growth response to elevated CO₂, suggesting thatplasticity in root allocation is relatively unimportant in determininggrowth response. Average N uptake rate at elevated comparedto ambient CO₂was decreased by 21–29%. In contrast, elevatedCO₂increased PNUE by approx. 50% due to a corresponding risein the CO₂-saturation factor for carboxylation at elevated CO₂.The model predicted that the decreased N uptake rate at elevatedCO₂lowered RGR modestly, but this effect was counterbalancedby an increase in PNUE resulting in a positive CO₂effect ongrowth. Increased PNUE may also explain why in many experimentselevated CO₂enhances biomass accumulation despite a significantdrop in tissue nitrogen concentration. The formulation of theFB model as presented here successfully predicted plant growthresponses to elevated CO₂. It also proved effective in resolvingwhich plant properties had the greatest leverage on such responses.Copyright 2000 Annals of Botany Company Elevated CO₂, functional balance model, Helianthus annuus L., N uptake, photosynthetic nitrogen use efficiency, root:shoot ratio     

Responses to CO2 Enrichment by Two Genotypes of Arabidopsis thaliana Differing in their Sensitivity to Nutrient Availability

The responses of two genotypes of Arabidopsis thaliana, whichdiffer in their sensitivities to nutrients to present and predictedfuture CO₂ concentration were determined under rich vs. poornutrient regimes on the basis of both single traits and thewhole plant. Based on individual traits, the two genotypes respondedsimilarly to CO₂ enrichment for all the traits measured exceptfor rate of increase in crown diameter, for which a decreasewas observed in the less nutrient-sensitive genotype grown atincreased CO₂. Based on the overall response of the whole plant,by analysing groups of plant traits using multivariate analysis,the two genotypes differed substantially from one another andboth responded more strongly to nutrient availability than toCO₂ concentration, especially for traits measured at harvestthat related to reproductive fitness. The less nutrient-sensitivegenotype also showed a weaker overall response to CO₂, and thepattern of the overall response was strikingly similar at differentnutrient supply. In contrast, the more nutrient-sensitive genotyperesponded more strongly to CO₂ than the less nutrient-sensitivegenotype, and responded differently to CO₂ at low vs. high nutrientavailability.Copyright 1995, 1999 Academic Press Plasticity, CO₂ enrichment, nutrient status, nutrient x CO₂ interaction, Arabidopsis thaliana, canonical analysis     

Direct and Maternal Effects of Elevated CO2on Early Root Growth of GerminatingArabidopsis thalianaSeedlings

Individuals ofArabidopsis thaliana, collected in different naturalpopulations, were grown in controlled and elevated CO₂in a glasshouse.Following germination, root growth of progeny of different linesof these populations was studied in control and elevated atmosphericCO₂. No significant direct effect of atmospheric CO₂concentrationcould be demonstrated on root growth. An important parentaleffect was apparent, namely that root length and branching weredecreased in seeds collected from a mother plant which had beengrown in elevated CO₂. This was correlated with smaller seeds,containing less nitrogen. These parental effects were geneticallyvariable. We conclude that CO₂may affect plant fitness via parentaleffects on seed size and early root growth and that the geneticvariability shown in our study demonstrates thatArabidopsispopulationswill evolve in the face of this new selective pressure.Copyright1998 Annals of Botany Company Root growth, root branching, seed, elevated CO₂, natural population,Arabidopsis thaliana, parental effect.     

Stomatal Characteristics of Four Native Herbs Following Exposure to Elevated CO2

Contrasting effects on the stomatal index (SI), stomatal density,epidermal cell size and number were observed in four chalk grasslandherbs (Sanguisorba minor Scop., Lotus corniculatus L., Anthyllisvulneraria L. and Plantago media L.) following exposure to elevatedcarbon dioxide concentrations (CO₂) in controlled environmentgrowth cabinets. SI of S. minor increased for both leaf surfaces,whilst in A. vulneraria and P. media SI decreased on one surfaceonly. In L. corniculatus , no differences in SI were observedas epidermal cell density changed in parallel with stomataldensity. In L. corniculatus and S. minor stomatal density increasedon both surfaces, whereas in P. media it decreased; in A. vulnerariastomatal density decreased on the abaxial leaf surface alonefollowing exposure to elevated CO₂. In the latter three species,SI changed because stomatal density did not change in parallelwith epidermal cell density. The results suggest elevated CO₂is either directly or indirectly affecting cell differentiationand thus stomatal initiation in the meristem. In S. minor and P. media leaf growth increased in elevated CO₂,because of increased cell expansion of epidermal cells, whereasin L. corniculatus, epidermal cell size decreased and greaterleaf growth was because of an increase in epidermal cell divisions.In A. vulneraria, leaf size did not change, but increased cellexpansion on the adaxial surface suggests CO₂ affects leaf surfacesdifferently, either directly or indirectly at the cell differentiationstage or as the leaf grows. These results suggest component species of a plant communitymay differ in their response to elevated CO₂. Predicting theeffect of environmental change is therefore difficult.Copyright1994, 1999 Academic Press Elevated CO₂, Sanguisorba minor (salad burnet), Lotus corniculatus (birdsfoot trefoil), Anthyllis vulneraria (kidney vetch), Plantago media (hoary plantain), stomatal index, stomatal density, epidermal cell size     

A Model for the Interaction of Low Temperature, ABA, IAA, and CO2 in the Control of Stomatal Behaviour

In the chilling sensitive (C.S.) species Phaseolus vulgarisit was found that at 22 ?C ABA induced stomatal closure butthis effect was dependent on the presence of CO₂. In the absenceof CO₂ the effect of ABA was completely lost. In contrast toABA, the effect of IAA at 22 ?C was to increase stomatal openingas the IAA concentration increased from 10^–2 to 10 molm^–3, and this effect was dependent upon the presence ofCO₂. However, at 5 ?C the action of ABA was reversed and itwas found to induce stomatal opening when fed via the transpirationstream in excised leaves. Similarly, the CO₂ response characteristicswere reversed at low temperatures as removal of CO₂ from theatmosphere caused stomatal closure. However, the effect of IAAat 5 ?C in the presence of CO₂ and with or without ABA was toincrease stomatal aperture with increasing IAA concentration.Significantly, ABA was found to have no effect upon aperturein the presence of CO₂ when IAA was added. The interactive effectsof ABA, IAA, CO₂ and low temperature are discussed in relationto a model proposed by the authors. Key words: IAA, ABA, CO₂, Stomata     

Effect of chronically elevated CO2 on CA1 neuronal excitability

To study the effect of chronically elevated CO₂ on the excitability and function of neurons, we exposed mice to 7.5–8% CO₂ for 2 wk (starting at 2 days of age) and examined the properties of freshly dissociated hippocampal neurons. Neurons from control mice (CON) and from mice exposed to chronically elevated CO₂ had similar resting membrane potentials and input resistances. CO₂-exposed neurons, however, had a lower rheobase and a higher Na⁺ current density (580 ± 73 pA/pF; n = 27 neurons studied) than did CON neurons (280 ± 51 pA/pF, n = 34; P < 0.01). In addition, the conductance-voltage curve was shifted in a more negative direction in CO₂-exposed than in CON neurons (midpoint of the curve was –46 ± 3 mV for CO₂ exposed and –34 ± 3 mV for CON, P < 0.01), while the steady-state inactivation curve was shifted in a more positive direction in CO₂-exposed than in CON neurons (midpoint of the curve was –59 ± 2 mV for CO₂ exposed and –68 ± 3 mV for CON, P < 0.01). The time constant for deactivation at –100 mV was much smaller in CO₂-exposed than in CON neurons (0.8 ± 0.1 ms for CO₂ exposed and 1.9 ± 0.3 ms for CON, P < 0.01). Immunoblotting for Na⁺ channel proteins (subtypes I, II, and III) was performed on the hippocampus. Our data indicate that Na⁺ channel subtype I, rather than subtype II or III, was significantly increased (43%, n = 4; P < 0.05) in the hippocampi of CO₂-exposed mice. We conclude that in mice exposed to elevated CO₂, 1) increased neuronal excitability is due to alterations in Na⁺ current and Na⁺ channel characteristics, and 2) the upregulation of Na⁺ channel subtype I contributes, at least in part, to the increase in Na⁺ current density. sodium ion channels; oxygen deprivation     

Contrasting CO2 and temperature effects on leaf growth of perennial ryegrass in spring and summer

The effects of increased atmospheric carbon dioxide (CO₂) of700 µmol mol^–1 and increased air temperature of+ 4C were examined in Lolium perenne L. cv. Vigor, growingin semi-controlled greenhouses. Leaf growth, segmental elongationrates (SER), water relations, cell wall (tensiometric) extensibility(%P) and epidermal cell lengths (ECL) were measured in expandingleaves in spring and summer. In elevated CO₂, shoot dry weight (SDW) increased in mid-summer.In both seasons, SDW decreased in elevated air temperatureswith this reduction being greater in summer as compared to spring.Specific leaf area (SLA) decreased in elevated CO₂ and in CO₂ temperature in both seasons. In spring, increased leaf extensionand SER in elevated CO₂ were linked with increased ECL, %P andfinal leaf size whilst in summer all were reduced. In high temperature,leaf extension, SER, %P and final leaf size were reduced inboth seasons. In elevated CO₂ temperature, leaf extension,SER, %P, and ECL increased in spring, but final leaf size remainedunaltered, whilst in summer all decreased. Mid-morning waterpotential did not differ with CO₂ or temperature treatments.Leaf turgor pressure increased in elevated CO₂ in spring andremained similar to the control in summer whilst solute potentialdecreased in spring and increased in summer. Contrasting seasonalgrowth responses of L. perenne in response to elevated CO₂ andtemperature suggests pasture management may change in the future.The grazing season may be prolonged, but whole season productivitymay become more variable than today. Key words: Lolium perenne, ryegrass, CO₂ and temperature, leaf extension, cell wall rheology     

Hypoxia, temperature, and pH/CO2 effects on respiratory discharge from a turtle brain stem preparation

Johnson, Stephen M., Rebecca A. Johnson, and Gordon S. Mitchell. Hypoxia, temperature, andpH/CO₂ effects on respiratory discharge from a turtle brain stem preparation. J. Appl. Physiol. 84(2): 649-660, 1998.An in vitrobrain stem preparation from adult turtles (Chrysemyspicta) was used to examine the effects of anoxia andincreased temperature and pH/CO₂on respiration-related motor output. At pH ~7.45, hypoglossal (XII)nerve roots produced patterns of rhythmic bursts (peaks) of discharge(0.74 ± 0.07 peaks/min, 10.0 ± 0.6 s duration) that werequantitatively similar to literature reports of respiratory activity inconscious, vagotomized turtles. Respiratory discharge was stable for 6 h at 22°C; at 32°C, peak amplitude and frequency progressivelyand reversibly decreased with time. Two hours of hypoxia had no effecton respiratory discharge. Acutely increasing bath temperature from 22 to 32°C decreased episode and peak duration and increased peakfrequency. Changes in pH/CO₂increased peak frequency from zero at pH 8.00-8.10 to maxima of0.81 ± 0.01 and 1.44 ± 0.02 peaks/min at 22°C (pH 7.32) and32°C (pH 7.46), respectively;pH/CO₂ sensitivity was similar atboth temperatures. We conclude that1) insensitivity to hypoxiaindicates that rhythmic discharge does not reflect gasping behavior,2) increased temperature altersrespiratory discharge, and 3)central pH/CO₂ sensitivity isunaffected by temperature in this preparation (i.e.,Q₁₀ ~1.0).