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Ventilatory responses to respiratory and metabolic acid-base disturbances in cats
Respiration Physiology (1987) 67, 69-83
69
Elsevier
RSP 01235
Ventilatory responses to respiratory and metabolic acid-base disturbances in cats
J.J. Schuitmaker, A. Berkenbosch, J. DeGoede and C.N. Olievier Department of Physiology and Physiological Physics, University of Leiden, Leiden, The Netherlands
(Accepted for publication 26 September 1986)
Abstract. To determine the relative importance of the peripheral and central chemoreceptors in the ventilatory response to acute metabolic a c i d - b a s e disturbances we measured the normoxic ventilatory response to acute respiratory and metabolic acidosis and alkalosis in 10 chloralose-urethane anesthetized cats using a technique of vertebral artery perfusion that allows one to independently manipulate the Paco2, Pao2 and the H + concentration of the blood in the systemic circulation (peripheral) and the blood perfusing the brain stem (central) (Berkenbosch et al., 1979). The ventilation could be satisfactorily described by a linear function of the peripheral and central arterial H ÷ concentration and the central P a c o 2. Mean values ( + SEM) found for the peripheral arterial H ÷ sensitivity and the isocapnic central arterial H ÷ sensitivity were 26.0 _+ 3.2 and 12.7 + 1.8 ml. min - 1. nM - 1, respectively; the isohydric central arterial CO 2 sensitivity was 545.9 + 96.7 ml. m i n - 1. k P a - 1 We conclude that in the ventilatory response to an acute metabolic a c i d - b a s e disturbance both the peripheral and central chemoreceptors play a role. However, the sensitivity of the peripheral chemoreceptors to isocapnic changes in the arterial H ÷ concentration is twice as large as the sensitivity of the central chemoreceptors. It is argued that in the adaptation of the ventilation to an acute metabolic acidosis the stimulatory effect of the peripheral chemoreceptors is counteracted by a diminished stimulation of the central chemoreceptors. A c i d - b a s e balance; Central chemoreceptor; Control of breathing; Metabolic acidosis; Peripheral chemoreceptors; Ventilation
The mechanisms underlying the ventilatory adaptation to an acute metabolic acid-base disturbance are still not well understood. Especially the relative importance of central and peripheral chemoreceptors is a subject of considerable controversy. According to Mitchell and Singer (1965) the adaptive ventilatory response to metabolic acid-base disturbances is initiated and maintained by the peripheral chemoreceptors. On the other hand, Fencl et aL (1966) advocate that the peripheral chemoreceptors do not play an Correspondence address: J.J. Schuitmaker, Department of Physiology and Physiological Physics, University of Leiden, Wassenaarseweg 62, P.O. Box 9604, 2300 RC Leiden, The Netherlands. 0034-5687/87/$03.50 © 1987 Elsevier Science Publishers B.V. (Biomedical Division)
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J.J. SCHUITMAKER et al.
important role during the ventilatory adaptation to a chronic metabolic acid-base disturbance. Experimental data supporting the theory of Mitchell (Nattie, 1983) or the theory of Fencl (Kaehny and Jackson, 1979; Steinbrook e t a l . , 1984) have been published. The commonly used technique in these experiments for separating the effects of the peripheral and central chemoreceptors in the ventilatory adaptation to acute metabolic acid-base disturbances has been denervation of the peripheral chemoreceptors. However, the interpretation of the results of denervation experiments rests on the assumption that the removal of a set of chemoreceptors removes only their own specific influence on ventilation (Dempsey and Forster, 1982). A promising technique, designed to quantitatively separate peripheral and central ventilatory effects of arterial H + concentration and CO2 tension without necessitating denervation is a technique in which the brain stem is artificially perfused with the cat's own blood. With this technique, which is extensively described by Berkenbosch et al. (1979) the CO2 and 0 2 tensions and the pH of the blood perfusing the brain stem and of the blood in the systemic circulation can be manipulated independently. Recently, using this technique, we confirmed that the ventilatory response of the peripheral chemoreceptors to metabolic or respiratory acid-base disturbances depends only on the arterial H + concentration (Schuitmaker et al., 1986). It is the purpose of this paper to gain more insight into the role played by the peripheral and central chemoreceptors during the ventilatory adaptation to acute metabolic acid-base disturbances by separating their effects on ventilation by artificially perfusing the brain stem. The main result of our study is that the ventilatory response to an acute metabolic acid-base disturbance is mediated by both the peripheral and central chemosensitive structures. The hypothesis of Fencl et al. (1966) that there is no role for the peripheral chemoreceptors in the adaptation of the ventilation to metabolic acid-base disturbances is not tenable, at least in anesthetized cats.
Materials and methods Technique. Experiments were performed on 10 cats of either sex, weighing 3.1-4.0 kg using a technique of artificial perfusion of the brain stem. The essence of this technique is to perfuse the brain stem of anesthetized cats with blood in which the blood gas tensions (called Paco2C and p cao2) can be varied independently from the blood gas tensions in the systemic circulation (called PaPo2 and PAP2). To this end blood is drawn from a femoral artery, led through a gas exchanger where desired blood gas tensions are imposed and subsequently infused into a cannulated vertebral artery at an infusion rate of 6-7 ml. rain ~. The other vertebral artery is clamped. This infusion rate has been shown to be sufficient to cause an ample overflow into the circle of Willis under conditions where there is cerebral vasodilatation (Berkenbosch et al., 1979; Beek et al., 1984). Acid-base disturbances were induced in the systemic circulation as well as in the central circulation. In three experiments we induced acid-base disturbances exclu-
VENTILATORY RESPONSES TO ACID-BASE DISTURBANCES
71
sively in the blood perfusing the brain stem. To this end the original technique was extended with a mixing chamber located in a shunt of the extracorporeal circuit in front of the gas exchanger (Schuitmaker et al., 1986). By infusion of small amounts of 0.3 M HC1 or 0.6 M N a H C O 3 into the mixing chamber the H + concentration of the blood perfusing the brain stem ( [H + ]~) could be changed. The H + concentration of the blood in the systemic circulation ( [H + ]ap) was kept constant in these experiments by infusion of HC1 or N a H C O 3 into the vena cava. Using the technique of artificial perfusion of the brain stem together with this mixing chamber enabled us to manipulate independently the 02 and CO2 tensions and the p H in the two artificially separated circulatory systems. Anesthesia was induced with 15 m g . k g - 1 ketamine hydrochloride intramuscularly, followed by inhalation of a gas mixture containing 0.8-1.2~o halothane and 30~o Oz in N 2. After cannulation of the right femoral vein 20 m g . k g - t chloralose and 100 m g . k g ~ urethane were slowly injected while at the same time the halothane concentration in the inspirate was reduced to zero. About one hour later infusion of a chloralose-urethane solution was started (1 m g . k g ~ . h - ~ chloralose and 5 mg. kg ~ • h ~urethane). Via the cannulated left femoral vein a polyethylene catheter was inserted into the vena cava for the administration of an HC1 or a NaHCO3 solution. The trachea was cannulated and connected to an open-ended tubing through which a flow of gas was delivered from a gas mixing system in excess of the inspiratory demand. The gas mixing system consists of three mass flow controllers (Type AFC-260, Advanced Semiconductor Materials) by which the flows of pure 02, N2 and CO 2 could be set at a desired rate. A microcomputer ( P D P 11/23) delivered the steering signal for the mass flow controllers to adjust the composition of the inspiratory gas mixture so that the end-tidal CO z and O 2 tensions could be set at any desired level. Temperature was monitored by a rectal thermistor and maintained within 1 ° C in the range from 36.4 to 38.3 °C by a heating pad and an infrared lamp. Right femoral arterial blood pressure was measured by a strain gauge. Details about the surgical procedures are given by Berkenbosch et al. (1979). Inspiratory and expiratory flow were measured using a Fleisch no. 0 pneumotachograph connected to a differential pressure transducer (Statham). The flow signal was electronically integrated to yield a volume signal. The CO2 concentration in the tracheal gas was measured with a fast infrared analyzer (Gould Godart M K 2 capnograph) and the O2 concentration with a fast zirconium oxide cell (Jaeger 0 2 Test). Carbon dioxide tensions in the blood of both circulatory systems were measured continuously with catheter type Pco2 electrodes (General Electric). Oxygen tensions were measured continuously with Clark-type electrodes mounted into a catheter (O.D. 1 mm). Blood pH was also measured continuously in both circulatory systems with combined glass-reference electrodes (Radiometer, type E 5037) connected to a pH meter (Philips, type PW 9409). The acid-base status of the animals was determined with a conventional sample method (Radiometer B M S 2 Mk2) in blood samples drawn at regular time intervals.
Measurements.
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J.J. SCHUITMAKER et al.
All signals were recorded on polygraphs, digitized (sample frequency 40 Hz) and processed by a PDP 11/23 micro computer. Moving averages over 20 breaths were calculated and stored every 15 sec for the respiratory timing, tidal volume, minute ventilation, arterial blood pressure, heart rate, perfusion pressure, rectal temperature, pH and gas tensions in blood, end-tidal and inspired gas. Details of measurements have been described previously (Berkenbosch et al., 1979; Schuitmaker et al., 1986).
EXPERIMENTAL DESIGN Experiments were performed during peripheral and central normoxia (Pa~2 and Pa~): about 15 kPa). There were two types of experimental protocol. The purpose of these experiments was to investigate whether the ventilatory effect of isohydric CO2 changes depends on the central acid-base level. To this end central steady-state ventilatory CO2 response curves were assessed during acute metabolic acid-base disturbances in the central circulation only. This was done by infusion of small amounts of 0.3 M HC1 or 0.6 M NaHCO3 with a dose of 3-6 mmol. h - 1 in the blood going to the brain stem with the aid of a mixing chamber. This centrally infused HC1 or NaHCO3 can reach the systemic circulation via the venous return. When the central induced acid-base disturbances are large it is not possible to keep the peripheral H + concentration at the original level by manipulating the CO 2 tension in the inspirate only. Therefore the acid-base status in the systemic circulation was kept constant by infusion of N a H C O 3 or HC1 into the vena cava. The PaPo~ and PaP~ were kept constant (coefficients of variation less than 3 To and 0.5 To, respectively) by adjusting the CO 2 and 02 concentrations in the inspirate. The PaSo 2 range covered was from 3.0 to 5.9 kPa; the central H + ion concentration ranged from 22 to 90 nM (pH 7.658-7.046). Protocol 1 : Central H ÷ and C O 2 responses.
The purpose of these experiments was to determine the peripheral H + and central isocapnic H + and isohydric CO2 sensitivities. Assuming that the steady-state ventilatory response of the peripheral chemoreceptors is a unique function of the peripheral arterial hydrogen ion concentration ([H+]P), an assumption for which there are strong indications (Schuitmaker e t a l . , 1986, and references cited therein), there is only one [H + ]~ sensitivity regardless whether the [H + ]P is changed by CO 2 or fixed acid. The mixing chamber was removed from the experimental set-up. The peripheral H + sensitivity at normal base excess was determined by manipulating the CO 2 concentration in the inspirate. Steady-state ventilation was measured at a normocapnic PaSo2 level of about 4 kPa. Subsequently an acute respiratory acidosis in the systemic circulation was induced by changing the PaSo2 in 2 or 3 steps of 0.5 to 1 kPa by augmenting the inspiratory CO 2 concentration. At each new level of [ H + ]~ the steady-state ventilation was measured. The Paco ~ and Pao~ in the central circulation were kept constant. The
Protocol 2: Peripheral H + and central H + and C 0 2 responses.
V E N T I L A T O R Y R E S P O N S E S TO A C I D - B A S E D I S T U R B A N C E S
73
peripheral Pao2 was kept constant by adjusting the inspired 02 concentration. The PaPo: range covered was from 3 to 10 kPa. After these measurements the acid-base status was changed in the peripheral and central circulation by rapid infusion of about 1 mmol of a 0.3 M HC1 or 0.6 M N a H C O 3 solution into the vena cava followed by a maintenance dose to keep the base excess level constant. The overall H + range was from 32.7 to 97.5 nM (pH 7.485-7.011). When ventilation had reached a steady-state, usually about 15 min after a change in base excess level, another peripheral steady-state ventilatory H + response curve was determined by changing the P a P o . This procedure was repeated two to four times. At least at one acid-base level a central CO 2 response curve was assessed by changing the Paco2.C The duration and the complexity of the experiments did not allow the assessment of the central CO 2 response curve at each base excess level. However, from the results of the experiments of protocol 1 it could be concluded that the central isohydric CO 2 sensitivity does not depend on the central acid-base level. Therefore the central isohydric CO 2 sensitivity can be reasonably assessed by one central CO 2 response curve. Note that this protocol is essentially based on the assumption that the steady-state ventilatory response of the peripheral chemoreceptors to respiratory or metabolic acid-base disturbances is a unique function of the arterial H + concentration for which, as remarked above, there is good evidence.
Data analysis.
The ventilation (VE) measured in the experiments in which the acid-base disturbances were induced only centrally, keeping the PaPo2, PaP 2 and [H ÷ ]ap constant (protocol 1) was fitted with the function of the [H + ]~ and Paco2-C. VE = S h [H
+
]aC + So° Paco2C - K1
(1)
in which S h represents the central isocapnic H ÷ sensitivity (called from now on central H + sensitivity) and S~ the central isohydric CO2 sensitivity. The parameter K~ is a constant which depends on peripheral conditions. The measured ventilation in protocol 2 was fitted with the function of the [H + ][, [H + ]c and PaSo2:
VE= Sh[H +]ap+ Sh [H +l~+ S~Pa~o ~ - K
(2)
The parameter Sph in this equation represents the peripheral H + sensitivity and the parameter K is a constant.
Results In three cats the central steady-state ventilatory C O 2 response curve was determined at several central base excess levels during constant peripheral conditions (protocol 1). The results of a typical experiment are shown in fig. 1. The steady-state ventilation is plotted against the central H + at four different central base excess levels in which the
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J.J. S C H U I T M A K E R et al.
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[ H+]ac (nmol I-1) Fig. l. Ventilation (VE) as a function of the central arterial H + concentration. Plotted are 4 central ventilatory response lines in which the [H +]2 was changed by altering the Pa~-,o2 (open symbols) and 1 central isocapnic ventilatory H + response line (closed symbols). Note that the isocapnic ventilatory H * response line is constructed out of measured points belonging to the different ventilatory response lines. See text for further details.
[H + ].~ is changed by altering the central Paco2. It illustrates that changing the central base excess level causes a displacement of the central steady-state ventilatory CO 2 response curve without a manifest change in slope. Also one central isocapnic ventilatory H + response curve is shown illustrating the presence of a central ventilatory H ~ effect. It can be seen that the stimulatory effect of raising the H + by increasing the p ca c o ~ on ventilation is much greater than the stimulatory effect of increasing the [H ~ ]~; by fixed acid. The same phenomena were observed in the other cats. We emphasize that the ventilatory response curves to changes in the Paco2C contain both the effects of [H + ]~ and PaSo ~proper. For each cat values were determined for the central isocapnic H + sensitivity S h, the central isohydric CO2 sensitivity S c and K~ at every central base excess level with multiple regression analysis using eq. (1). The results are presented in table 1. The values in each cat were not significantly different using two-way analysis of variance (P values for S h, S~ and K~ are 0.7, 0.9 and 0.9, respectively) indicating that they are not influenced by the central base excess level. Figure 2 shows the ventilatory reaction ('¢E) of a cat, which was subjected to protocol 2 after a rapid infusion of 0.6 M N a H C O 3 into the vena cava followed by a maintenance dose. The infusion causes a fast change in the peripheral arterial pH followed by a fast decrease in ventilation which must be due to the peripheral chemoreceptors as the pH change has not reached the perfused brain stem yet. After a time delay caused by the
V E N T I L A T O R Y R E S P O N S E S TO A C I D - B A S E D I S T U R B A N C E S
75
TABLE 1 Normoxic values for the ventilatory sensitivity S~ ( m l . m i n l . n M 1) t o [H+].~ (riM), S~: (ml" min ~ ' k P a - ~) to PaSo 2 (kPa) and for the constant K~ (ml. rain l) at four different central base excess levels as indicated by the bicarbonate concentration (mM) in the blood perfusing the brain stem at the lowest P a c o 2 level. Exp.
521
no.
S~
S~
K~
[HCO 3 ]
1
32.8 32.0 35.3 29.5 32.4 1.2
1278.6 985.9 843.2 1153.6 1065.3 95.3
4945.8 3706.8 3276.2 4245.7 4043.6 360.2
9.9 22.6 31.2 15.2
1 2 3 4
27.8 31.0 23.5 28.5 27.7 1.6
542.4 352.9 399.6 418.9 428.5 40.4
2716.7 1864.3 1809.3 2163.5 2138.5 207.9
17.3 13.6 12.1 16.5
1 2 3 4
8.3 9.9 9.5 7.8 8.9 0.5
735.7 659.7 906.2 462.7 691.1 91.9
1844.0 1686.5 2694.2 672.5 1724.3 4t4.6
16.2 13.8 9.2 6.8
2 3 4 mean SEM 523
mean SEM 525
mean SEM
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76
J.J. S C H U I T M A K E R et al.
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experimental set-up a change in the central arterial p H occurs which is somewhat slower due to the mixing of the systemic blood with the blood in the reservoir of the gas exchanger. This causes a further decrease in ventilation which in all probability is of central origin. This figure also illustrates that after an acute change in base excess a steady-state ventilation is reached after c a . 15 min. In fig. 3 steady-state ventilation is plotted against the peripheral arterial H + concentration at three different central base excess levels. The lower and middle ventilatory response curves were obtained by changing the [H + ]P via changes in the PaPo2. The upper response curve, however, was obtained by i.v. infusion of 0.3 molar HC1 into the vena cava under isocapnic conditions. These results illustrate that the peripheral H + sensitivity does not depend on the central base excess level. This implies that we can write K l in eq. (1) as
K, = - S ~ [H ÷]~ + K thus arriving at eq. (2). The individual data of the eight cats subjected to protocol 2 were fitted with eq. (2) using multiple regression analysis. The estimated parameters are presented in table 2. The mean value ( + S E M ) estimated for the central isohydric CO 2 sensitivity S~, was 545.9 + 96.7 ml- min 1. kPa 1. The mean values estimated for the central isocapnic H ÷ sensitivity S h and for the peripheral H ÷ sensitivity S h were 12.7 + 1.8 and 26.0 + 3.2 ml- m i n - 1. nM 1, respectively. It is of considerable interest to note that the isocapnic peripheral H + sensitivity is about twice the central isocapnic H + sensitivity.
VENTILATORY RESPONSES TO ACID-BASE DISTURBANCES
77
TABLE 2 Normoxic values for the ventilatory sensitivity S~, (ml.min 1.nM-I) to [H+]ap (nM), S) (ml" rain ~ ~•uM - ~) to [H + ]~ (nM) and S~ (ml. rain - i. kPa - i ) to Pa~.o~ (kPa) and for the constant K (ml.min 1). Exp.
S~
S~
S~
K
312 512 513 515 517 519 524 525
39.8 27.6 30.5 14.7 15.6 17.2 29.5 33.0
9.1 20.2 21.1 9.3 8.3 10.5 9.4 13.3
244.5 645.3 628.7 315.2 333.4 435.2 1081.7 683.3
1155.2 4268.0 4329.7 1102.0 2063.5 2710.0 4898.4 4158.4
mean SEM
26.0 3.2
12.7 1.8
545.9 96.7
3085.7 538.3
Discussion The technique o f artificially perfusing the medulla, pons and cerebellum with the cat's own blood enables one to m a n i p u l a t e the Pco2, Po2 and the p H o f the blood perfusing the central chemosensitive structures i n d e p e n d e n t from those in the systemic circulation in which the peripheral c h e m o r e c e p t o r s are located. In this way a physical separation o f the effects of central and peripheral chemical stimuli on ventilation can be accomplished without the necessity o f denervating a n d vagotomising the cat. In the present study we used this technique to assess quantitatively the sensitivity, in terms of ventilation, of the peripheral and central c h e m o r e c e p t o r s to acute a c i d - b a s e disturbances. This means that our results are only valid for ventilatory effects of a c i d - b a s e disturbances for time periods o f a few hours (cf. D e m p s e y and Forster, 1982). It was found that the peripheral H + sensitivity was on the average twice the central isocapnic H + sensitivity. W e described ventilation as a linear function of the [H ÷ ]~P, [H + ]~ and P a ~ , o : In fig. 4, upper panel, the ventilation predicted by eq. (2) is plotted vs the m e a s u r e d ventilation for a representative experiment. In fig. 4, lower panel, the residuals are plotted vs the time o f m e a s u r e m e n t of the same experiment. They both illustrate that the ventilation can be adequately described with eq. (2). However, a change in central P a c o 2 will be a c c o m p a n i e d by a change in central [H + ]a introducing a correlation between the i n d e p e n d e n t variables Paco2C and [H + ]X. This so-called multicollinearity will give rise to unstable estimates o f the regression coefficients S~ and S~. To avoid this the [ H + ]~ was also altered, independently o f the P a S o : by fixed acid. As a result the multicollinearity in our experiments was found to be small since the variance inflation was less than 2 in m o s t cases and never exceeded 3 (see Belsley et al., 1980).
78
J.J. SCHUITMAKER et al. EXP.519 I
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time Fig. 4. Upper panel: Predicted ventilation plotted against the measured ventilation using eq. (2) of a representative experiment. Drawn is the line of identity. Lower panel: Same experiment. Residuals plotted against the time of measurement illustrating that during an experiment no manifest trend develops in the residuals.
VENTILATORY RESPONSES TO ACID-BASE DISTURBANCES
79
We have determined the sensitivities of the peripheral and central chemoreceptors to acute acid-base disturbances in terms of ventilation. To compare the contribution of changes in the [H + ]a and P a c o 2 on ventilation we have to know how much a change in Pa¢o 2 alters the arterial H ÷ ion concentration. From our data, using log P a c o 2 vs pH a graphs it could be inferred that a change in the P a c o 2 from 4 to 5 kPa under normal acid-base conditions of the cat (a base excess of about - 7.2 mM) causes an increase of the [H + ]a of 5.8 nM ( e l Borison et al., 1977). Using this value for the increase in [H + ]a in eq. (2) it then follows that the peripheral chemoreceptors contribute 20~o to the change in ventilation during an acute respiratory acid-base disturbance. The central chemoreceptors contribute 80Yo to the change in ventilation, 70~o through their sensitivity to P a c o 2 and 10~o through their sensitivity to [H + ]a. In several studies the ventilatory response to changes in H + ion concentration and the Pco2 in the arterial blood has been investigated (Domizi et al., 1959; Saito et al., 1960; Natsui, 1970; Kuwana and Natsui, 1981). It was found in these studies that the change in ventilation caused by a respiratory acid-base disturbance can be attributed for approximately 70Yo to the change in P a c o 2 and for 30yo to the change in the H + ion concentration. The magnitude of the P a c o ~effect on ventilation is in agreement with our findings. Assuming that under steady-state conditions CO2 without changing the [H + ]a does not stimulate the peripheral chemoreceptors (Schuitmaker et al., 1986) the P a c o ~effect is of central origin. The above mentioned studies, however, did not separate the H + effect into a central and peripheral part. Our results show that this H + effect is for 2/3 of peripheral and 1/3 of central origin. In a study concerning the effects of the "arterial [H + ] on the Pco2 at which apnea occurs Kuwana and N atsui (1981) calculated the relative contributions of the arterial [H +] and Pco2 in stimulating the peripheral and central chemoreceptors. They concluded that the H + sensitivity of the peripheral chemoreceptors is about equal to the H + sensitivity of the central chemoreceptors. Our results suggest, however, that the H + sensitivity of the peripheral chemoreceptors is about two times as large as the H + sensitivity of the central chemoreceptors. These different experimental results of Kuwana and Natsui could be caused by their experimental protocol which deviated considerably from ours. They made use of artificially ventilated, carotid body denervated vagotomized cats. Our cats were intact and spontaneously breathing. From data presented by Teppema et al. (1983) it follows that the ratio of the slope of the ventilatory response line of the [ H + ] in the medullary surface extracellular fluid ([H + ]ECF) obtained by changing the end-tidal Pco2 and the slope of the VQEv s [H ÷ ]ECF response line obtained by isocapnically changing the [H + ]a is about 3. Assuming that an acute metabolic isocapnic [H + ]~ change is for approximately 30~o reflected in a concomitant change of [H + ]ECF (Ahmad et aL, 1976) and that a step change in end-tidal Pco~ is for about 75~o reflected in a change in CO 2 tension of the ECF (see Olievier et al., 1982) it follows from our data that this ratio is ca. 2.5. This fair agreement suggests that the sensitivities by measuring [H + ] changes of the ECF on the medullary surface and those found when measuring [ H + ] changes in arterial blood are compatible. Equation (2) is only valid for H + concentrations and CO2 tensions measured in the
80
J.J. SCHUITMAKER et al.
arterial blood. According to this equation the sensitivity of the central chemoreceptors is separated into a central (isocapnic) H + sensitivity and a central isohydric CO 2 sensitivity. This does not necessarily imply that these agents exert separate effects on the central chemosensitive structures proper. However, experimental evidence is accumulating that CO2 and H + in the E C F do exert separate effects on the central chemosensitive structures. Teppema et al. (1983), in a study concerning the effects of respiratory and metabolic acid-base disturbances on medullary extracellular fluid pH found strong indications that both Pco2 and [H + ] in the ECF exert independent ventilatory responses via the central chemoreceptors. Similar results were obtained by Eldridge et al. (1985), who concluded, measuring phrenic nerve activity responses to hypercapnic and metabolically induced acidosis in the ECF of cats, that the [H + ]~<:~, does not represent a unique stimulus to the central chemoreceptors. Also Shams (1985), measuring the pH and Pco2 on the ventral surface of the medulla in cats came to the conclusion that CO2 as well as H + exert independent effects on respiration via the central chemoreceptors. If the central chemoreceptors do have separate responses to H + and CO2 in the steady-state they differ in this aspect from the peripheral chemoreceptors where the H + ion is probably the unique stimulus (Schuitmaker et al., 1986, and references cited therein). There is no consistency in the existing literature concerning the specific role of the peripheral arterial chemoreceptors in the ventilatory adaptation to acute acid-base disturbances (Dempsey and Forster, 1982). According to Mitchell and Singer (1965) the peripheral chemoreceptors initiate and maintain the adaptive ventilatory response to metabolic acid-base disturbances. Contrary to this hypothesis Fencl et al. (1966) conclude that resting ventilation is a single function of [H + ] in the cerebral interstitial fluid during all degrees of a chronic metabolic acid-base disturbance. Experimental data in support of the view of Mitchell or Fencl have both been published. Kaehny and Jackson (1979) working with unanesthetized dogs of which the carotid sinus nerves were sectioned and Javaheri e t al. (1979) in anesthetized dogs both concluded from their data that the peripheral chemoreceptors are not essential for the ventilatory adaptation to metabolic acidosis. Also Steinbrook et al. (1984) found in goats that a respiratory adaptation to an acute metabolic acidosis occurs after ablation of the carotid bodies. On the other hand Bainton (1978) found in dogs that the ventilatory response to an increase of the [H + ]a was lost after carotid body denervation. Nattie (1983) found in awake rabbits which were carotid body denervated an immediate ventilatory response to i.v. infusion of HC1. This response was approximately 1/3 of the response observed in intact rabbits and he concluded that the peripheral chemoreceptors accounted for about 2/3 of the ventilatory response to metabolic acidosis. Our results show that in the ventilatory response to an acute metabolic acid-base disturbance both the peripheral and central chemoreceptors play a role. It is instructive to calculate the contribution to the increase in ventilation by the central and peripheral chemoreceptors following an increase in the arterial [H + ], for the case in which the CO2 concentration in the inspirate is zero. It follows from eq. (A3) in the appendix that the increase in ventilation is a balanced effect of the peripheral and central H + sensitivities
VENTILATORY RESPONSES TO ACID-BASE DISTURBANCES
81
a n d the i s o h y d r i c central C O 2 sensitivity. T a k i n g for the m e t a b o l i c C O 2 p r o d u c t i o n 25 m l . rain 1, for the b a r o m e t r i c p r e s s u r e 100 k P a a n d a P a c o 2 o f 4 k P a it c a n be c a l c u l a t e d f r o m the v a l u e s o f S~, S h a n d S~ (table 2) that the net c o n t r i b u t i o n o f the central c h e m o r e c e p t o r s is always n e g a t i v e in an a c u t e m e t a b o l i c acidosis. H e n c e the s t i m u l a t o r y effect on v e n t i l a t i o n by the p e r i p h e r a l c h e m o r e c e p t o r s is c o u n t e r a c t e d by a d e p r e s s a n t effect o f the c e n t r a l o n e s as the d e c r e a s e in P a c o 2 c a u s e s a d i m i n i s h e d c e n t r a l stimulation w h i c h is larger t h a n the c e n t r a l s t i m u l a t i o n by H + . A f t e r peripheral c h e m o d e n e r v a t i o n v e n t i l a t i o n still i n c r e a s e s after an a c u t e m e t a b o l i c acidosis, but n o w due to the central s t i m u l a t o r y effect o f H + . T h i s i n c r e a s e is a b o u t 1/3 o f that o f the n o n - d e n e r v a t e d cat since Sp/S~ h h is a b o u t 2. T h i s c a l c u l a t i o n is c o n s i s t e n t with the result f o u n d by N a t t i e (1983) in rabbits. H o w e v e r , the c o n c l u s i o n o f N a t t i e t h a t the p e r i p h e r a l c h e m o r e c e p t o r s a c c o u n t e d for 2/3 o f the v e n t i l a t o r y r e s p o n s e to m e t a b o l i c a c i d o s i s m a y be w r o n g since, in a n o n - d e n e r v a t e d animal, there m a y be a c e n t r a l d e p r e s s i o n o f v e n t i l a t i o n d u r i n g a c u t e a c i d o s i s w h e n the C O 2 c o n c e n t r a t i o n in the i n s p i r a t e is zero. S i m i l a r c a l c u l a t i o n s c a n be p e r f o r m e d for m i l d alkalosis.
In conclusion, o u r study in cats s h o w s t h a t the v e n t i l a t o r y r e s p o n s e to an a c u t e m e t a b o l i c a c i d - b a s e d i s t u r b a n c e is m e d i a t e d by b o t h the p e r i p h e r a l a n d central chemoreceptors.
F u r t h e r m o r e , it s h o w s t h a t the v e n t i l a t o r y H + sensitivity o f the
peripheral c h e m o r e c e p t o r s is a b o u t t w o t i m e s as large as the i s o c a p n i c H + sensitivity o f the c e n t r a l c h e m o r e c e p t o r s .
Acknowledgements. This study was subsidized by the Foundation for Medical Research FUNGO: grant no. 900-519-043. We are indebted to Mr. L. Philips for his skillful work in the surgical preparation of the animals and to Mr. M.M. Rancuret for technical assistance.
Appendix We found that the ventilation during normoxia could be adequately described by eq. (2) VE - S~ [H+] p + Sh [H+]~ + S~ PaSo 2 - K When the brain stem is not artificially perfused [H + ]P = [H ÷ ]~ = [H ÷ ]a and PaSo 2 = Paco 2 so that we can write VE = (Shy + S~)[H+]a + S~ Paco 2 - K
(al)
For the metabolic hyperbola we take VE - (PB" Qco2)/(Paco2 - PIco2) + ~/D
(A2)
in which PB is the barometric pressure, ~'co2 the metabolic CO 2 production, "V'Dthe dead space ventilation and Plco 2 the CO 2 pressure of the inspirate. Linearizing eq. (A1) and (A2) around a certain PaSo 2 located at the intersection of eq. (AI) and (A2) we find for small but fixed changes in [H + ]~ (A [H + ]a) the following relation between the change in ventilation (AVE) and A[H + ]a AVE = (S~, + S~)[1 - S~/{S~ + (PB. 9co2).(Pa~::o2 - PIco2) 2}]A[n+]a
(A3)
82
J.J. S C H U I T M A K E R et al.
using h APaco 2 = [-(Sp, + Sc)/{S cc + (P8' Vco2) ' . (Paco2 o - Plco2)- 2}]A[H + ]a
Equation (A3) describes the change in ventilation for small changes in [H + ]a. It thus enables one to estimate the contributions of the peripheral and central chemoreceptors during acute metabolic acid-base disturbances when the inspiratory Pco2 is kept constant instead of the arterial Pco2. It can be shown that the central contribution to the change in ventilation during acidosis is negative if c h Sph . Sc/S c > (P8.9co2).(Pa~:o2 - Pico2)
2.
It is zero if c Sph . Sc/S ch = (P8" Vco2).(Pa~:o2 - PIco2) -2
and positive if h
c~
h
Sp" So/Sc < (PB" Vco2) " (PaSo2 - PIco2) - 2 For alkalosis the reverse inequality signs hold true.
References Ahmad, H.R., J. Berndt and H.H. Loeschcke (1976). Bicarbonate exchange between blood, brain extracellular fluid and brain cells at maintained pCO 2. In: A c i d - B a s e Homeostasis of the Brain Extracellular Fluid and the Respiratory Control System, edited by H.H. Loeschcke. Stuttgart, Thieme, pp. 19-27. Bainton, C. R. (1978). Canine ventilation after a c i d - b a s e infusions, exercise and carotid body denervation. J. Appl. Physiol. 44: 28-35. Beek, J . H . G . M . van, A. Berkenbosch, J. DeGoede and C.N. Olievier (1984). Effects of brain stem hypoxaemia on the regulation of breathing. Respir. Physiol. 57: 171-188. Belsley, D.A., E. Kuh and R. E. Welsch (1980). Regression Diagnostics. New York, John Wiley and Sons. Berkenbosch, A., J. Heeringa, C.N. Olievier and E.W. Kruyt (1979). Artificial perfusion of the pontomedullarly region of cats. A method for separation of central and peripheral effects of chemical stimulation on ventilation. Respir. Physiol. 37: 347-364. Borison, H. L., J. H. Hurst, L. E. McCarthy and R. Rosenstein (1977). Arterial hydrogen ion versus CO~ on depth and rate of breathing in decerebrate cats. Respir. Physiol. 30:311-325. Dempsey, J. A. and H.V. Forster (1982). Mediation of ventilatory adaptations. Physiol. Rev. 62: 262-346. Domizi, D.B., J.F. Perkins, Jr., and J.S. Byrne (1959). Ventilatory response to fixed acid evaluated by "iso-Pco 2' technique. J. Appl. Physiol. 14: 557-561. Eldridge, F.L., J.P. Kiley and D.E. Millhorn (1985). Respiratory responses to medullary hydrogen ion changes in cats: different effects of respiratory and metabolic acidosis. J. Physiol. (London) 358: 285-297. Fencl, V., T. B. Miller and J.R. Pappenheimer (1966). Studies on the respiratory response to disturbances of a c i d - b a s e balance, with deductions concerning the ionic composition of cerebral interstitial fluid. Am. J. Physiol. 210: 459-472. Javaheri, S., L. Herrera and H. Kazemi (1979). Ventilatory drive in acute metabolic acidosis. J. Appl. Physiol. 45: 913-918. Kaehny, W.D. and J.T. Jackson (1979). Respiratory response to HCI acidosis in dogs after carotid body denervation. J. Appl. Physiol. 46:1138-1142. Kuwana, S. and T. Natsui (1981). Effect of arterial [H +] on threshold Pco2 of the respiratory system in vagotomized and carotid sinus nerve denervated cats..I. Physiol. (London) 318: 223-237.
VENTILATORY RESPONSES TO A C I D - B A S E DISTURBANCES
83
Mitchell, R.A. and M.M. Singer (1965). Respiration and cerebrospinal fluid pH in metabolic acidosis and alkalosis. J. Appl. Physiol. 20:905-911. Natsui, T. (1970). Respiratory response to arterial H ÷ at different levels of arterial Pco2 during hyperoxia or hypoxia. Pfliigers Arch. 316: 34-50. Nattie, E. E. (1983). Ventilation during acute HCI infusion in intact and chemodenervated conscious rabbits. Respir. Physiol. 54: 97-107. Olievier, C.N., A. Berkenbosch, J.H.G.M. van Beek, J. DeGoede and Ph.H. Quanjer (1982). Hypoxia, cerebrospinal fluid Pco2 and central depression of ventilation. Bull. Eur. Physiopathol. Respir. 18: 165-172. Saito, K., Y. Honda and N. Hasumura (1960). Evaluation of respiratory response to changes in Pco2 and hydrogen ion concentration of arterial blood in rabbits and dogs. Jpn. J. Physiol. 10: 634-645. Schuitmaker, J.J., A. Berkenbosch, J. DeGoede and C.N. Olievier (1986). Effects of CO 2 and H ÷ on the ventilatory response to peripheral chemoreceptor stimulation. Respir. Physiol. 64: 69-79. Shams, H. (1985). Differential effects of CO 2 and H + as central stimuli of respiration in the cat. d. Appl. Physiol. 58: 357-364. Steinbrook, R.A., S. Javaheri, R.A. Gabel, J. C. Donovan, D.E. Leith and V. Fencl (1984). Respiration of chemodenervated goats in acute metabolic acidosis. Respir. Physiol. 56: 51-60. Teppema, L.J., P.W.J.A. Barts, H.Th. Folgering and J.A.M. Evers (1983). Effects of respiratory and (isocapnic) metabolic arterial acid-base disturbances on medullary extracellular fluid pH and ventilation in cats. Respir. Physiol. 53: 375-379.
69
Elsevier
RSP 01235
Ventilatory responses to respiratory and metabolic acid-base disturbances in cats
J.J. Schuitmaker, A. Berkenbosch, J. DeGoede and C.N. Olievier Department of Physiology and Physiological Physics, University of Leiden, Leiden, The Netherlands
(Accepted for publication 26 September 1986)
Abstract. To determine the relative importance of the peripheral and central chemoreceptors in the ventilatory response to acute metabolic a c i d - b a s e disturbances we measured the normoxic ventilatory response to acute respiratory and metabolic acidosis and alkalosis in 10 chloralose-urethane anesthetized cats using a technique of vertebral artery perfusion that allows one to independently manipulate the Paco2, Pao2 and the H + concentration of the blood in the systemic circulation (peripheral) and the blood perfusing the brain stem (central) (Berkenbosch et al., 1979). The ventilation could be satisfactorily described by a linear function of the peripheral and central arterial H ÷ concentration and the central P a c o 2. Mean values ( + SEM) found for the peripheral arterial H ÷ sensitivity and the isocapnic central arterial H ÷ sensitivity were 26.0 _+ 3.2 and 12.7 + 1.8 ml. min - 1. nM - 1, respectively; the isohydric central arterial CO 2 sensitivity was 545.9 + 96.7 ml. m i n - 1. k P a - 1 We conclude that in the ventilatory response to an acute metabolic a c i d - b a s e disturbance both the peripheral and central chemoreceptors play a role. However, the sensitivity of the peripheral chemoreceptors to isocapnic changes in the arterial H ÷ concentration is twice as large as the sensitivity of the central chemoreceptors. It is argued that in the adaptation of the ventilation to an acute metabolic acidosis the stimulatory effect of the peripheral chemoreceptors is counteracted by a diminished stimulation of the central chemoreceptors. A c i d - b a s e balance; Central chemoreceptor; Control of breathing; Metabolic acidosis; Peripheral chemoreceptors; Ventilation
The mechanisms underlying the ventilatory adaptation to an acute metabolic acid-base disturbance are still not well understood. Especially the relative importance of central and peripheral chemoreceptors is a subject of considerable controversy. According to Mitchell and Singer (1965) the adaptive ventilatory response to metabolic acid-base disturbances is initiated and maintained by the peripheral chemoreceptors. On the other hand, Fencl et aL (1966) advocate that the peripheral chemoreceptors do not play an Correspondence address: J.J. Schuitmaker, Department of Physiology and Physiological Physics, University of Leiden, Wassenaarseweg 62, P.O. Box 9604, 2300 RC Leiden, The Netherlands. 0034-5687/87/$03.50 © 1987 Elsevier Science Publishers B.V. (Biomedical Division)
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J.J. SCHUITMAKER et al.
important role during the ventilatory adaptation to a chronic metabolic acid-base disturbance. Experimental data supporting the theory of Mitchell (Nattie, 1983) or the theory of Fencl (Kaehny and Jackson, 1979; Steinbrook e t a l . , 1984) have been published. The commonly used technique in these experiments for separating the effects of the peripheral and central chemoreceptors in the ventilatory adaptation to acute metabolic acid-base disturbances has been denervation of the peripheral chemoreceptors. However, the interpretation of the results of denervation experiments rests on the assumption that the removal of a set of chemoreceptors removes only their own specific influence on ventilation (Dempsey and Forster, 1982). A promising technique, designed to quantitatively separate peripheral and central ventilatory effects of arterial H + concentration and CO2 tension without necessitating denervation is a technique in which the brain stem is artificially perfused with the cat's own blood. With this technique, which is extensively described by Berkenbosch et al. (1979) the CO2 and 0 2 tensions and the pH of the blood perfusing the brain stem and of the blood in the systemic circulation can be manipulated independently. Recently, using this technique, we confirmed that the ventilatory response of the peripheral chemoreceptors to metabolic or respiratory acid-base disturbances depends only on the arterial H + concentration (Schuitmaker et al., 1986). It is the purpose of this paper to gain more insight into the role played by the peripheral and central chemoreceptors during the ventilatory adaptation to acute metabolic acid-base disturbances by separating their effects on ventilation by artificially perfusing the brain stem. The main result of our study is that the ventilatory response to an acute metabolic acid-base disturbance is mediated by both the peripheral and central chemosensitive structures. The hypothesis of Fencl et al. (1966) that there is no role for the peripheral chemoreceptors in the adaptation of the ventilation to metabolic acid-base disturbances is not tenable, at least in anesthetized cats.
Materials and methods Technique. Experiments were performed on 10 cats of either sex, weighing 3.1-4.0 kg using a technique of artificial perfusion of the brain stem. The essence of this technique is to perfuse the brain stem of anesthetized cats with blood in which the blood gas tensions (called Paco2C and p cao2) can be varied independently from the blood gas tensions in the systemic circulation (called PaPo2 and PAP2). To this end blood is drawn from a femoral artery, led through a gas exchanger where desired blood gas tensions are imposed and subsequently infused into a cannulated vertebral artery at an infusion rate of 6-7 ml. rain ~. The other vertebral artery is clamped. This infusion rate has been shown to be sufficient to cause an ample overflow into the circle of Willis under conditions where there is cerebral vasodilatation (Berkenbosch et al., 1979; Beek et al., 1984). Acid-base disturbances were induced in the systemic circulation as well as in the central circulation. In three experiments we induced acid-base disturbances exclu-
VENTILATORY RESPONSES TO ACID-BASE DISTURBANCES
71
sively in the blood perfusing the brain stem. To this end the original technique was extended with a mixing chamber located in a shunt of the extracorporeal circuit in front of the gas exchanger (Schuitmaker et al., 1986). By infusion of small amounts of 0.3 M HC1 or 0.6 M N a H C O 3 into the mixing chamber the H + concentration of the blood perfusing the brain stem ( [H + ]~) could be changed. The H + concentration of the blood in the systemic circulation ( [H + ]ap) was kept constant in these experiments by infusion of HC1 or N a H C O 3 into the vena cava. Using the technique of artificial perfusion of the brain stem together with this mixing chamber enabled us to manipulate independently the 02 and CO2 tensions and the p H in the two artificially separated circulatory systems. Anesthesia was induced with 15 m g . k g - 1 ketamine hydrochloride intramuscularly, followed by inhalation of a gas mixture containing 0.8-1.2~o halothane and 30~o Oz in N 2. After cannulation of the right femoral vein 20 m g . k g - t chloralose and 100 m g . k g ~ urethane were slowly injected while at the same time the halothane concentration in the inspirate was reduced to zero. About one hour later infusion of a chloralose-urethane solution was started (1 m g . k g ~ . h - ~ chloralose and 5 mg. kg ~ • h ~urethane). Via the cannulated left femoral vein a polyethylene catheter was inserted into the vena cava for the administration of an HC1 or a NaHCO3 solution. The trachea was cannulated and connected to an open-ended tubing through which a flow of gas was delivered from a gas mixing system in excess of the inspiratory demand. The gas mixing system consists of three mass flow controllers (Type AFC-260, Advanced Semiconductor Materials) by which the flows of pure 02, N2 and CO 2 could be set at a desired rate. A microcomputer ( P D P 11/23) delivered the steering signal for the mass flow controllers to adjust the composition of the inspiratory gas mixture so that the end-tidal CO z and O 2 tensions could be set at any desired level. Temperature was monitored by a rectal thermistor and maintained within 1 ° C in the range from 36.4 to 38.3 °C by a heating pad and an infrared lamp. Right femoral arterial blood pressure was measured by a strain gauge. Details about the surgical procedures are given by Berkenbosch et al. (1979). Inspiratory and expiratory flow were measured using a Fleisch no. 0 pneumotachograph connected to a differential pressure transducer (Statham). The flow signal was electronically integrated to yield a volume signal. The CO2 concentration in the tracheal gas was measured with a fast infrared analyzer (Gould Godart M K 2 capnograph) and the O2 concentration with a fast zirconium oxide cell (Jaeger 0 2 Test). Carbon dioxide tensions in the blood of both circulatory systems were measured continuously with catheter type Pco2 electrodes (General Electric). Oxygen tensions were measured continuously with Clark-type electrodes mounted into a catheter (O.D. 1 mm). Blood pH was also measured continuously in both circulatory systems with combined glass-reference electrodes (Radiometer, type E 5037) connected to a pH meter (Philips, type PW 9409). The acid-base status of the animals was determined with a conventional sample method (Radiometer B M S 2 Mk2) in blood samples drawn at regular time intervals.
Measurements.
72
J.J. SCHUITMAKER et al.
All signals were recorded on polygraphs, digitized (sample frequency 40 Hz) and processed by a PDP 11/23 micro computer. Moving averages over 20 breaths were calculated and stored every 15 sec for the respiratory timing, tidal volume, minute ventilation, arterial blood pressure, heart rate, perfusion pressure, rectal temperature, pH and gas tensions in blood, end-tidal and inspired gas. Details of measurements have been described previously (Berkenbosch et al., 1979; Schuitmaker et al., 1986).
EXPERIMENTAL DESIGN Experiments were performed during peripheral and central normoxia (Pa~2 and Pa~): about 15 kPa). There were two types of experimental protocol. The purpose of these experiments was to investigate whether the ventilatory effect of isohydric CO2 changes depends on the central acid-base level. To this end central steady-state ventilatory CO2 response curves were assessed during acute metabolic acid-base disturbances in the central circulation only. This was done by infusion of small amounts of 0.3 M HC1 or 0.6 M NaHCO3 with a dose of 3-6 mmol. h - 1 in the blood going to the brain stem with the aid of a mixing chamber. This centrally infused HC1 or NaHCO3 can reach the systemic circulation via the venous return. When the central induced acid-base disturbances are large it is not possible to keep the peripheral H + concentration at the original level by manipulating the CO 2 tension in the inspirate only. Therefore the acid-base status in the systemic circulation was kept constant by infusion of N a H C O 3 or HC1 into the vena cava. The PaPo~ and PaP~ were kept constant (coefficients of variation less than 3 To and 0.5 To, respectively) by adjusting the CO 2 and 02 concentrations in the inspirate. The PaSo 2 range covered was from 3.0 to 5.9 kPa; the central H + ion concentration ranged from 22 to 90 nM (pH 7.658-7.046). Protocol 1 : Central H ÷ and C O 2 responses.
The purpose of these experiments was to determine the peripheral H + and central isocapnic H + and isohydric CO2 sensitivities. Assuming that the steady-state ventilatory response of the peripheral chemoreceptors is a unique function of the peripheral arterial hydrogen ion concentration ([H+]P), an assumption for which there are strong indications (Schuitmaker e t a l . , 1986, and references cited therein), there is only one [H + ]~ sensitivity regardless whether the [H + ]P is changed by CO 2 or fixed acid. The mixing chamber was removed from the experimental set-up. The peripheral H + sensitivity at normal base excess was determined by manipulating the CO 2 concentration in the inspirate. Steady-state ventilation was measured at a normocapnic PaSo2 level of about 4 kPa. Subsequently an acute respiratory acidosis in the systemic circulation was induced by changing the PaSo2 in 2 or 3 steps of 0.5 to 1 kPa by augmenting the inspiratory CO 2 concentration. At each new level of [ H + ]~ the steady-state ventilation was measured. The Paco ~ and Pao~ in the central circulation were kept constant. The
Protocol 2: Peripheral H + and central H + and C 0 2 responses.
V E N T I L A T O R Y R E S P O N S E S TO A C I D - B A S E D I S T U R B A N C E S
73
peripheral Pao2 was kept constant by adjusting the inspired 02 concentration. The PaPo: range covered was from 3 to 10 kPa. After these measurements the acid-base status was changed in the peripheral and central circulation by rapid infusion of about 1 mmol of a 0.3 M HC1 or 0.6 M N a H C O 3 solution into the vena cava followed by a maintenance dose to keep the base excess level constant. The overall H + range was from 32.7 to 97.5 nM (pH 7.485-7.011). When ventilation had reached a steady-state, usually about 15 min after a change in base excess level, another peripheral steady-state ventilatory H + response curve was determined by changing the P a P o . This procedure was repeated two to four times. At least at one acid-base level a central CO 2 response curve was assessed by changing the Paco2.C The duration and the complexity of the experiments did not allow the assessment of the central CO 2 response curve at each base excess level. However, from the results of the experiments of protocol 1 it could be concluded that the central isohydric CO 2 sensitivity does not depend on the central acid-base level. Therefore the central isohydric CO 2 sensitivity can be reasonably assessed by one central CO 2 response curve. Note that this protocol is essentially based on the assumption that the steady-state ventilatory response of the peripheral chemoreceptors to respiratory or metabolic acid-base disturbances is a unique function of the arterial H + concentration for which, as remarked above, there is good evidence.
Data analysis.
The ventilation (VE) measured in the experiments in which the acid-base disturbances were induced only centrally, keeping the PaPo2, PaP 2 and [H ÷ ]ap constant (protocol 1) was fitted with the function of the [H + ]~ and Paco2-C. VE = S h [H
+
]aC + So° Paco2C - K1
(1)
in which S h represents the central isocapnic H ÷ sensitivity (called from now on central H + sensitivity) and S~ the central isohydric CO2 sensitivity. The parameter K~ is a constant which depends on peripheral conditions. The measured ventilation in protocol 2 was fitted with the function of the [H + ][, [H + ]c and PaSo2:
VE= Sh[H +]ap+ Sh [H +l~+ S~Pa~o ~ - K
(2)
The parameter Sph in this equation represents the peripheral H + sensitivity and the parameter K is a constant.
Results In three cats the central steady-state ventilatory C O 2 response curve was determined at several central base excess levels during constant peripheral conditions (protocol 1). The results of a typical experiment are shown in fig. 1. The steady-state ventilation is plotted against the central H + at four different central base excess levels in which the
74
J.J. S C H U I T M A K E R et al.
EXP521 I
I
I
!
I
3
&A E
1
i
t
40
--!
/
~0
I
80
[ H+]ac (nmol I-1) Fig. l. Ventilation (VE) as a function of the central arterial H + concentration. Plotted are 4 central ventilatory response lines in which the [H +]2 was changed by altering the Pa~-,o2 (open symbols) and 1 central isocapnic ventilatory H + response line (closed symbols). Note that the isocapnic ventilatory H * response line is constructed out of measured points belonging to the different ventilatory response lines. See text for further details.
[H + ].~ is changed by altering the central Paco2. It illustrates that changing the central base excess level causes a displacement of the central steady-state ventilatory CO 2 response curve without a manifest change in slope. Also one central isocapnic ventilatory H + response curve is shown illustrating the presence of a central ventilatory H ~ effect. It can be seen that the stimulatory effect of raising the H + by increasing the p ca c o ~ on ventilation is much greater than the stimulatory effect of increasing the [H ~ ]~; by fixed acid. The same phenomena were observed in the other cats. We emphasize that the ventilatory response curves to changes in the Paco2C contain both the effects of [H + ]~ and PaSo ~proper. For each cat values were determined for the central isocapnic H + sensitivity S h, the central isohydric CO2 sensitivity S c and K~ at every central base excess level with multiple regression analysis using eq. (1). The results are presented in table 1. The values in each cat were not significantly different using two-way analysis of variance (P values for S h, S~ and K~ are 0.7, 0.9 and 0.9, respectively) indicating that they are not influenced by the central base excess level. Figure 2 shows the ventilatory reaction ('¢E) of a cat, which was subjected to protocol 2 after a rapid infusion of 0.6 M N a H C O 3 into the vena cava followed by a maintenance dose. The infusion causes a fast change in the peripheral arterial pH followed by a fast decrease in ventilation which must be due to the peripheral chemoreceptors as the pH change has not reached the perfused brain stem yet. After a time delay caused by the
V E N T I L A T O R Y R E S P O N S E S TO A C I D - B A S E D I S T U R B A N C E S
75
TABLE 1 Normoxic values for the ventilatory sensitivity S~ ( m l . m i n l . n M 1) t o [H+].~ (riM), S~: (ml" min ~ ' k P a - ~) to PaSo 2 (kPa) and for the constant K~ (ml. rain l) at four different central base excess levels as indicated by the bicarbonate concentration (mM) in the blood perfusing the brain stem at the lowest P a c o 2 level. Exp.
521
no.
S~
S~
K~
[HCO 3 ]
1
32.8 32.0 35.3 29.5 32.4 1.2
1278.6 985.9 843.2 1153.6 1065.3 95.3
4945.8 3706.8 3276.2 4245.7 4043.6 360.2
9.9 22.6 31.2 15.2
1 2 3 4
27.8 31.0 23.5 28.5 27.7 1.6
542.4 352.9 399.6 418.9 428.5 40.4
2716.7 1864.3 1809.3 2163.5 2138.5 207.9
17.3 13.6 12.1 16.5
1 2 3 4
8.3 9.9 9.5 7.8 8.9 0.5
735.7 659.7 906.2 462.7 691.1 91.9
1844.0 1686.5 2694.2 672.5 1724.3 4t4.6
16.2 13.8 9.2 6.8
2 3 4 mean SEM 523
mean SEM 525
mean SEM
EXP519
1 phi; / __
--
,/
f/
j '
70
PHI"
l--
" -
J'/'
70~
{h rain ~)1
-" --
OJ'~T~T7
0
f
-
,
r r r v ~ T T r T r T r r l r r T i
10
20
tlmO
( r
Fig. 2. Ventilatory reaction (?E) foUowing a rapid change in base excess by i.v. infusionofNaHCO3 (arrow). This causes a fast change in the peripheral arterial pH ( p H i ) and in ventilation. After a time delay caused by the experimental set-up, a relatively slow change in the central arterial pH (pHi) and a further decrease in ventilation occurs. A steady-state ventilation is reached after about 15 min.
76
J.J. S C H U I T M A K E R et al.
225-
EXP 519 --
~ -
I
I
I
- - + - -
+--
4- --
~
I
I E
125
025
40
i
5b
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6'o - " - - 7 b
'
~o
~
90
[ H+]Pn mol I '1 Fig. 3. Steady-state ventilation (~'E) plotted against the peripheral arterial H + concentration ([H +]p) at three different central base excess levels. The lower and middle ventilatory response curves were obtained by changing the [H + ]P via changes in the PaPo2. The upper response curve was obtained by i.v. infusion of fixed acid. Note the similarity in responses. For further details see the text.
experimental set-up a change in the central arterial p H occurs which is somewhat slower due to the mixing of the systemic blood with the blood in the reservoir of the gas exchanger. This causes a further decrease in ventilation which in all probability is of central origin. This figure also illustrates that after an acute change in base excess a steady-state ventilation is reached after c a . 15 min. In fig. 3 steady-state ventilation is plotted against the peripheral arterial H + concentration at three different central base excess levels. The lower and middle ventilatory response curves were obtained by changing the [H + ]P via changes in the PaPo2. The upper response curve, however, was obtained by i.v. infusion of 0.3 molar HC1 into the vena cava under isocapnic conditions. These results illustrate that the peripheral H + sensitivity does not depend on the central base excess level. This implies that we can write K l in eq. (1) as
K, = - S ~ [H ÷]~ + K thus arriving at eq. (2). The individual data of the eight cats subjected to protocol 2 were fitted with eq. (2) using multiple regression analysis. The estimated parameters are presented in table 2. The mean value ( + S E M ) estimated for the central isohydric CO 2 sensitivity S~, was 545.9 + 96.7 ml- min 1. kPa 1. The mean values estimated for the central isocapnic H ÷ sensitivity S h and for the peripheral H ÷ sensitivity S h were 12.7 + 1.8 and 26.0 + 3.2 ml- m i n - 1. nM 1, respectively. It is of considerable interest to note that the isocapnic peripheral H + sensitivity is about twice the central isocapnic H + sensitivity.
VENTILATORY RESPONSES TO ACID-BASE DISTURBANCES
77
TABLE 2 Normoxic values for the ventilatory sensitivity S~, (ml.min 1.nM-I) to [H+]ap (nM), S) (ml" rain ~ ~•uM - ~) to [H + ]~ (nM) and S~ (ml. rain - i. kPa - i ) to Pa~.o~ (kPa) and for the constant K (ml.min 1). Exp.
S~
S~
S~
K
312 512 513 515 517 519 524 525
39.8 27.6 30.5 14.7 15.6 17.2 29.5 33.0
9.1 20.2 21.1 9.3 8.3 10.5 9.4 13.3
244.5 645.3 628.7 315.2 333.4 435.2 1081.7 683.3
1155.2 4268.0 4329.7 1102.0 2063.5 2710.0 4898.4 4158.4
mean SEM
26.0 3.2
12.7 1.8
545.9 96.7
3085.7 538.3
Discussion The technique o f artificially perfusing the medulla, pons and cerebellum with the cat's own blood enables one to m a n i p u l a t e the Pco2, Po2 and the p H o f the blood perfusing the central chemosensitive structures i n d e p e n d e n t from those in the systemic circulation in which the peripheral c h e m o r e c e p t o r s are located. In this way a physical separation o f the effects of central and peripheral chemical stimuli on ventilation can be accomplished without the necessity o f denervating a n d vagotomising the cat. In the present study we used this technique to assess quantitatively the sensitivity, in terms of ventilation, of the peripheral and central c h e m o r e c e p t o r s to acute a c i d - b a s e disturbances. This means that our results are only valid for ventilatory effects of a c i d - b a s e disturbances for time periods o f a few hours (cf. D e m p s e y and Forster, 1982). It was found that the peripheral H + sensitivity was on the average twice the central isocapnic H + sensitivity. W e described ventilation as a linear function of the [H ÷ ]~P, [H + ]~ and P a ~ , o : In fig. 4, upper panel, the ventilation predicted by eq. (2) is plotted vs the m e a s u r e d ventilation for a representative experiment. In fig. 4, lower panel, the residuals are plotted vs the time o f m e a s u r e m e n t of the same experiment. They both illustrate that the ventilation can be adequately described with eq. (2). However, a change in central P a c o 2 will be a c c o m p a n i e d by a change in central [H + ]a introducing a correlation between the i n d e p e n d e n t variables Paco2C and [H + ]X. This so-called multicollinearity will give rise to unstable estimates o f the regression coefficients S~ and S~. To avoid this the [ H + ]~ was also altered, independently o f the P a S o : by fixed acid. As a result the multicollinearity in our experiments was found to be small since the variance inflation was less than 2 in m o s t cases and never exceeded 3 (see Belsley et al., 1980).
78
J.J. SCHUITMAKER et al. EXP.519 I
I
I
I
t
I
I
I
I
I
I
I
I
I
[]
^
D
~E 1.
0
I
I
I
0
I
I
2
VE ~I,min-l>measured
EXP.519 I
I
I
I
I
I
I
I
I
I
I
I
0..~ 13 D D
o
[] ~ D
==
Dn[]
DD
m-o
q~
o
~-0.5
6
l
I
I
I
8
I
I
I
I
10
I
I
I
I
12
time
VENTILATORY RESPONSES TO ACID-BASE DISTURBANCES
79
We have determined the sensitivities of the peripheral and central chemoreceptors to acute acid-base disturbances in terms of ventilation. To compare the contribution of changes in the [H + ]a and P a c o 2 on ventilation we have to know how much a change in Pa¢o 2 alters the arterial H ÷ ion concentration. From our data, using log P a c o 2 vs pH a graphs it could be inferred that a change in the P a c o 2 from 4 to 5 kPa under normal acid-base conditions of the cat (a base excess of about - 7.2 mM) causes an increase of the [H + ]a of 5.8 nM ( e l Borison et al., 1977). Using this value for the increase in [H + ]a in eq. (2) it then follows that the peripheral chemoreceptors contribute 20~o to the change in ventilation during an acute respiratory acid-base disturbance. The central chemoreceptors contribute 80Yo to the change in ventilation, 70~o through their sensitivity to P a c o 2 and 10~o through their sensitivity to [H + ]a. In several studies the ventilatory response to changes in H + ion concentration and the Pco2 in the arterial blood has been investigated (Domizi et al., 1959; Saito et al., 1960; Natsui, 1970; Kuwana and Natsui, 1981). It was found in these studies that the change in ventilation caused by a respiratory acid-base disturbance can be attributed for approximately 70Yo to the change in P a c o 2 and for 30yo to the change in the H + ion concentration. The magnitude of the P a c o ~effect on ventilation is in agreement with our findings. Assuming that under steady-state conditions CO2 without changing the [H + ]a does not stimulate the peripheral chemoreceptors (Schuitmaker et al., 1986) the P a c o ~effect is of central origin. The above mentioned studies, however, did not separate the H + effect into a central and peripheral part. Our results show that this H + effect is for 2/3 of peripheral and 1/3 of central origin. In a study concerning the effects of the "arterial [H + ] on the Pco2 at which apnea occurs Kuwana and N atsui (1981) calculated the relative contributions of the arterial [H +] and Pco2 in stimulating the peripheral and central chemoreceptors. They concluded that the H + sensitivity of the peripheral chemoreceptors is about equal to the H + sensitivity of the central chemoreceptors. Our results suggest, however, that the H + sensitivity of the peripheral chemoreceptors is about two times as large as the H + sensitivity of the central chemoreceptors. These different experimental results of Kuwana and Natsui could be caused by their experimental protocol which deviated considerably from ours. They made use of artificially ventilated, carotid body denervated vagotomized cats. Our cats were intact and spontaneously breathing. From data presented by Teppema et al. (1983) it follows that the ratio of the slope of the ventilatory response line of the [ H + ] in the medullary surface extracellular fluid ([H + ]ECF) obtained by changing the end-tidal Pco2 and the slope of the VQEv s [H ÷ ]ECF response line obtained by isocapnically changing the [H + ]a is about 3. Assuming that an acute metabolic isocapnic [H + ]~ change is for approximately 30~o reflected in a concomitant change of [H + ]ECF (Ahmad et aL, 1976) and that a step change in end-tidal Pco~ is for about 75~o reflected in a change in CO 2 tension of the ECF (see Olievier et al., 1982) it follows from our data that this ratio is ca. 2.5. This fair agreement suggests that the sensitivities by measuring [H + ] changes of the ECF on the medullary surface and those found when measuring [ H + ] changes in arterial blood are compatible. Equation (2) is only valid for H + concentrations and CO2 tensions measured in the
80
J.J. SCHUITMAKER et al.
arterial blood. According to this equation the sensitivity of the central chemoreceptors is separated into a central (isocapnic) H + sensitivity and a central isohydric CO 2 sensitivity. This does not necessarily imply that these agents exert separate effects on the central chemosensitive structures proper. However, experimental evidence is accumulating that CO2 and H + in the E C F do exert separate effects on the central chemosensitive structures. Teppema et al. (1983), in a study concerning the effects of respiratory and metabolic acid-base disturbances on medullary extracellular fluid pH found strong indications that both Pco2 and [H + ] in the ECF exert independent ventilatory responses via the central chemoreceptors. Similar results were obtained by Eldridge et al. (1985), who concluded, measuring phrenic nerve activity responses to hypercapnic and metabolically induced acidosis in the ECF of cats, that the [H + ]~<:~, does not represent a unique stimulus to the central chemoreceptors. Also Shams (1985), measuring the pH and Pco2 on the ventral surface of the medulla in cats came to the conclusion that CO2 as well as H + exert independent effects on respiration via the central chemoreceptors. If the central chemoreceptors do have separate responses to H + and CO2 in the steady-state they differ in this aspect from the peripheral chemoreceptors where the H + ion is probably the unique stimulus (Schuitmaker et al., 1986, and references cited therein). There is no consistency in the existing literature concerning the specific role of the peripheral arterial chemoreceptors in the ventilatory adaptation to acute acid-base disturbances (Dempsey and Forster, 1982). According to Mitchell and Singer (1965) the peripheral chemoreceptors initiate and maintain the adaptive ventilatory response to metabolic acid-base disturbances. Contrary to this hypothesis Fencl et al. (1966) conclude that resting ventilation is a single function of [H + ] in the cerebral interstitial fluid during all degrees of a chronic metabolic acid-base disturbance. Experimental data in support of the view of Mitchell or Fencl have both been published. Kaehny and Jackson (1979) working with unanesthetized dogs of which the carotid sinus nerves were sectioned and Javaheri e t al. (1979) in anesthetized dogs both concluded from their data that the peripheral chemoreceptors are not essential for the ventilatory adaptation to metabolic acidosis. Also Steinbrook et al. (1984) found in goats that a respiratory adaptation to an acute metabolic acidosis occurs after ablation of the carotid bodies. On the other hand Bainton (1978) found in dogs that the ventilatory response to an increase of the [H + ]a was lost after carotid body denervation. Nattie (1983) found in awake rabbits which were carotid body denervated an immediate ventilatory response to i.v. infusion of HC1. This response was approximately 1/3 of the response observed in intact rabbits and he concluded that the peripheral chemoreceptors accounted for about 2/3 of the ventilatory response to metabolic acidosis. Our results show that in the ventilatory response to an acute metabolic acid-base disturbance both the peripheral and central chemoreceptors play a role. It is instructive to calculate the contribution to the increase in ventilation by the central and peripheral chemoreceptors following an increase in the arterial [H + ], for the case in which the CO2 concentration in the inspirate is zero. It follows from eq. (A3) in the appendix that the increase in ventilation is a balanced effect of the peripheral and central H + sensitivities
VENTILATORY RESPONSES TO ACID-BASE DISTURBANCES
81
a n d the i s o h y d r i c central C O 2 sensitivity. T a k i n g for the m e t a b o l i c C O 2 p r o d u c t i o n 25 m l . rain 1, for the b a r o m e t r i c p r e s s u r e 100 k P a a n d a P a c o 2 o f 4 k P a it c a n be c a l c u l a t e d f r o m the v a l u e s o f S~, S h a n d S~ (table 2) that the net c o n t r i b u t i o n o f the central c h e m o r e c e p t o r s is always n e g a t i v e in an a c u t e m e t a b o l i c acidosis. H e n c e the s t i m u l a t o r y effect on v e n t i l a t i o n by the p e r i p h e r a l c h e m o r e c e p t o r s is c o u n t e r a c t e d by a d e p r e s s a n t effect o f the c e n t r a l o n e s as the d e c r e a s e in P a c o 2 c a u s e s a d i m i n i s h e d c e n t r a l stimulation w h i c h is larger t h a n the c e n t r a l s t i m u l a t i o n by H + . A f t e r peripheral c h e m o d e n e r v a t i o n v e n t i l a t i o n still i n c r e a s e s after an a c u t e m e t a b o l i c acidosis, but n o w due to the central s t i m u l a t o r y effect o f H + . T h i s i n c r e a s e is a b o u t 1/3 o f that o f the n o n - d e n e r v a t e d cat since Sp/S~ h h is a b o u t 2. T h i s c a l c u l a t i o n is c o n s i s t e n t with the result f o u n d by N a t t i e (1983) in rabbits. H o w e v e r , the c o n c l u s i o n o f N a t t i e t h a t the p e r i p h e r a l c h e m o r e c e p t o r s a c c o u n t e d for 2/3 o f the v e n t i l a t o r y r e s p o n s e to m e t a b o l i c a c i d o s i s m a y be w r o n g since, in a n o n - d e n e r v a t e d animal, there m a y be a c e n t r a l d e p r e s s i o n o f v e n t i l a t i o n d u r i n g a c u t e a c i d o s i s w h e n the C O 2 c o n c e n t r a t i o n in the i n s p i r a t e is zero. S i m i l a r c a l c u l a t i o n s c a n be p e r f o r m e d for m i l d alkalosis.
In conclusion, o u r study in cats s h o w s t h a t the v e n t i l a t o r y r e s p o n s e to an a c u t e m e t a b o l i c a c i d - b a s e d i s t u r b a n c e is m e d i a t e d by b o t h the p e r i p h e r a l a n d central chemoreceptors.
F u r t h e r m o r e , it s h o w s t h a t the v e n t i l a t o r y H + sensitivity o f the
peripheral c h e m o r e c e p t o r s is a b o u t t w o t i m e s as large as the i s o c a p n i c H + sensitivity o f the c e n t r a l c h e m o r e c e p t o r s .
Acknowledgements. This study was subsidized by the Foundation for Medical Research FUNGO: grant no. 900-519-043. We are indebted to Mr. L. Philips for his skillful work in the surgical preparation of the animals and to Mr. M.M. Rancuret for technical assistance.
Appendix We found that the ventilation during normoxia could be adequately described by eq. (2) VE - S~ [H+] p + Sh [H+]~ + S~ PaSo 2 - K When the brain stem is not artificially perfused [H + ]P = [H ÷ ]~ = [H ÷ ]a and PaSo 2 = Paco 2 so that we can write VE = (Shy + S~)[H+]a + S~ Paco 2 - K
(al)
For the metabolic hyperbola we take VE - (PB" Qco2)/(Paco2 - PIco2) + ~/D
(A2)
in which PB is the barometric pressure, ~'co2 the metabolic CO 2 production, "V'Dthe dead space ventilation and Plco 2 the CO 2 pressure of the inspirate. Linearizing eq. (A1) and (A2) around a certain PaSo 2 located at the intersection of eq. (AI) and (A2) we find for small but fixed changes in [H + ]~ (A [H + ]a) the following relation between the change in ventilation (AVE) and A[H + ]a AVE = (S~, + S~)[1 - S~/{S~ + (PB. 9co2).(Pa~::o2 - PIco2) 2}]A[n+]a
(A3)
82
J.J. S C H U I T M A K E R et al.
using h APaco 2 = [-(Sp, + Sc)/{S cc + (P8' Vco2) ' . (Paco2 o - Plco2)- 2}]A[H + ]a
Equation (A3) describes the change in ventilation for small changes in [H + ]a. It thus enables one to estimate the contributions of the peripheral and central chemoreceptors during acute metabolic acid-base disturbances when the inspiratory Pco2 is kept constant instead of the arterial Pco2. It can be shown that the central contribution to the change in ventilation during acidosis is negative if c h Sph . Sc/S c > (P8.9co2).(Pa~:o2 - Pico2)
2.
It is zero if c Sph . Sc/S ch = (P8" Vco2).(Pa~:o2 - PIco2) -2
and positive if h
c~
h
Sp" So/Sc < (PB" Vco2) " (PaSo2 - PIco2) - 2 For alkalosis the reverse inequality signs hold true.
References Ahmad, H.R., J. Berndt and H.H. Loeschcke (1976). Bicarbonate exchange between blood, brain extracellular fluid and brain cells at maintained pCO 2. In: A c i d - B a s e Homeostasis of the Brain Extracellular Fluid and the Respiratory Control System, edited by H.H. Loeschcke. Stuttgart, Thieme, pp. 19-27. Bainton, C. R. (1978). Canine ventilation after a c i d - b a s e infusions, exercise and carotid body denervation. J. Appl. Physiol. 44: 28-35. Beek, J . H . G . M . van, A. Berkenbosch, J. DeGoede and C.N. Olievier (1984). Effects of brain stem hypoxaemia on the regulation of breathing. Respir. Physiol. 57: 171-188. Belsley, D.A., E. Kuh and R. E. Welsch (1980). Regression Diagnostics. New York, John Wiley and Sons. Berkenbosch, A., J. Heeringa, C.N. Olievier and E.W. Kruyt (1979). Artificial perfusion of the pontomedullarly region of cats. A method for separation of central and peripheral effects of chemical stimulation on ventilation. Respir. Physiol. 37: 347-364. Borison, H. L., J. H. Hurst, L. E. McCarthy and R. Rosenstein (1977). Arterial hydrogen ion versus CO~ on depth and rate of breathing in decerebrate cats. Respir. Physiol. 30:311-325. Dempsey, J. A. and H.V. Forster (1982). Mediation of ventilatory adaptations. Physiol. Rev. 62: 262-346. Domizi, D.B., J.F. Perkins, Jr., and J.S. Byrne (1959). Ventilatory response to fixed acid evaluated by "iso-Pco 2' technique. J. Appl. Physiol. 14: 557-561. Eldridge, F.L., J.P. Kiley and D.E. Millhorn (1985). Respiratory responses to medullary hydrogen ion changes in cats: different effects of respiratory and metabolic acidosis. J. Physiol. (London) 358: 285-297. Fencl, V., T. B. Miller and J.R. Pappenheimer (1966). Studies on the respiratory response to disturbances of a c i d - b a s e balance, with deductions concerning the ionic composition of cerebral interstitial fluid. Am. J. Physiol. 210: 459-472. Javaheri, S., L. Herrera and H. Kazemi (1979). Ventilatory drive in acute metabolic acidosis. J. Appl. Physiol. 45: 913-918. Kaehny, W.D. and J.T. Jackson (1979). Respiratory response to HCI acidosis in dogs after carotid body denervation. J. Appl. Physiol. 46:1138-1142. Kuwana, S. and T. Natsui (1981). Effect of arterial [H +] on threshold Pco2 of the respiratory system in vagotomized and carotid sinus nerve denervated cats..I. Physiol. (London) 318: 223-237.
VENTILATORY RESPONSES TO A C I D - B A S E DISTURBANCES
83
Mitchell, R.A. and M.M. Singer (1965). Respiration and cerebrospinal fluid pH in metabolic acidosis and alkalosis. J. Appl. Physiol. 20:905-911. Natsui, T. (1970). Respiratory response to arterial H ÷ at different levels of arterial Pco2 during hyperoxia or hypoxia. Pfliigers Arch. 316: 34-50. Nattie, E. E. (1983). Ventilation during acute HCI infusion in intact and chemodenervated conscious rabbits. Respir. Physiol. 54: 97-107. Olievier, C.N., A. Berkenbosch, J.H.G.M. van Beek, J. DeGoede and Ph.H. Quanjer (1982). Hypoxia, cerebrospinal fluid Pco2 and central depression of ventilation. Bull. Eur. Physiopathol. Respir. 18: 165-172. Saito, K., Y. Honda and N. Hasumura (1960). Evaluation of respiratory response to changes in Pco2 and hydrogen ion concentration of arterial blood in rabbits and dogs. Jpn. J. Physiol. 10: 634-645. Schuitmaker, J.J., A. Berkenbosch, J. DeGoede and C.N. Olievier (1986). Effects of CO 2 and H ÷ on the ventilatory response to peripheral chemoreceptor stimulation. Respir. Physiol. 64: 69-79. Shams, H. (1985). Differential effects of CO 2 and H + as central stimuli of respiration in the cat. d. Appl. Physiol. 58: 357-364. Steinbrook, R.A., S. Javaheri, R.A. Gabel, J. C. Donovan, D.E. Leith and V. Fencl (1984). Respiration of chemodenervated goats in acute metabolic acidosis. Respir. Physiol. 56: 51-60. Teppema, L.J., P.W.J.A. Barts, H.Th. Folgering and J.A.M. Evers (1983). Effects of respiratory and (isocapnic) metabolic arterial acid-base disturbances on medullary extracellular fluid pH and ventilation in cats. Respir. Physiol. 53: 375-379.