EFFECTS IN MAN OF ACUTE EXPERIMENTAL RESPIRATORY ALKALOSIS AND ACIDOSIS ON IONIC TRANSFERS IN THE TOTAL BODY FLUIDS

(1)

EFFECTS IN MAN OF ACUTE EXPERIMENTAL

RESPIRATORY ALKALOSIS AND ACIDOSIS ON

IONIC TRANSFERS IN THE TOTAL BODY

FLUIDS

J. R. Elkinton, … , E. S. Barker, J. K. Clark

J Clin Invest.

1955;

34(11)

:1671-1690.

https://doi.org/10.1172/JCI103221

.

Research Article

Find the latest version:

(2)

ALKALOSIS AND ACIDOSIS ON IONIC

TRANSFERS IN THE

TOTAL BODY FLUIDS

1

By J. R. ELKINTON,2 R. B. SINGER,3 E. S. BARKER,4 AND J. K. CLARK

(Fromthe Chemical SectionandRenal Sectionof theDepartment ofMedicine, theDepartment

of Research Medicine, and the Department of Biochemistry, the University of Pennsylvania School of Medicine, Philadelphia, Pa.)

(Submitted for publicationJanuary31, 1955; accepted August3, 1955)

Although the concept of tissue

buffering

was

formulated

early in the study of acid-base

dis-turbances, attention in the past has been directed

primarily towards the effects of these disturbances

on

blood and

extracellular fluid.

More

recently

the demonstration

by

Darrow

and

his co-workers

(1-3) of

exchanges of intracellular cations

during

clinical and

experimental potassium

deficiency

stimulated renewed interest in the

quantitative

aspects of the

buffering action of intracellular fluid.

Accordingly,

some

years ago the authors

under-took

quantitative

estimation of the effects in

man

of acute

experimental

respiratory

alkalosis and

acidosis

on

the components of the

multiphase

sys-tem

of red

cells,

plasma, interstitial

fluid,

and

"in-tracellular" fluid

(calculated

as

the

non-chloride

space),

as

well

as on

exchanges

in

the

kidney.

The latter renal effects will be

reported

in

another

paper

(4).

Our results indicate that in

man

the effects of

acute

hyperventilation

or

carbon

dioxide

inhala-tion

are

buffered

to a

large

extent

by

a

series of

linked ionic

exchanges

with

a

phase

or

phases

out-side the chloride space,

as

well

as

with buffer

sys-tems

within the blood and extracellular

fluid.

Since these

findings

were

reported

in abstract

(5-8)

similar conclusions have been reached

by

Giebisch, Berger,

and Pitts

(9),

on

the basis of

experimental respiratory

disturbances in

the

dog.

Estimates of the

importance

of tissue

buffering

1Laboratory facilities were aided by grants from the Life Insurance Medical Research Fund, the National Heart Institute of the United States Public Health Serv-ice (Grants H-340 and

H-405),

and the C. Mahlon Kline Fund for Development in the Department of Medicine.

2

_Established

_Investigator _of _the _American _Heart

Association.

3Present address: 501 Boylston Street, Boston, Mass.

4During tenure of Post-doctorate Fellowship of the National Institutes of Health, United States Public

HIealth

Service.

have also been made in acute metabolic alkalosis

and acidosis in man (10, 11) and in the dog

(12-14).

EXPERIMENTAL PROCEDURE AND METHODS

The effects of the two opposite types of respiratory

disturbance, hyperventilation and

_{CO_}

inhalation, were

assessed by comparison of the changes observed from the control period to the period during altered

respira-tion and from the latter to the recovery period. This assessment has been made by use of the balance technic;

the method ofcalculation is presented in the next section. Twelve normal men between the ages of 19 and 34, students and physicians, served as experimental subjects. All subjects werein the fasting state and the studies were

performed during the morning hours (8 A.M. to 12 noon); during this interval diurnal variations in acid-base balance and renal function should be small, uni-directional, and comparable between all experiments as

a control background. Standard loading of the sub-jects with water and solutes was carried out before and during the experiments as follows. In the respiratory alkalosis experiments, this consisted essentially in water loading only in amounts equivalent to urine excreted. In therespiratory acidosis experiments, Experiments 1 to

5 inclusive, the subjects were given 4.2 gm. NaHCO3 (50 mEq.) by mouthat -95 to - 150minutes to produce

a very slight metabolic alkalosis and ensure a neutral or alkaline urine before the onset of the stimulus; in the sixth experiment NaCl was given as a control. In all the latter experiments one liter of water was given with the salt as a priming solution to institute an adequate and relatively constant urine flow; subsequently water

was taken orally in amounts equivalent to urinary ex-cretion. All subjects remained recumbent throughout the experiments.

Following control periods of 47 to 74 minutes'

dura-tion, respiratory alkalosis was induced by voluntary hy-perventilation for approximately 30 minutes in 5 of the 6 experiments, and for twice that period in the last

ex-periment; respiratory acidosis was induced by breathing

7.5 to 7.7 per cent CO2 in air or 0, for periods of 21 to

30 minutes. Observations were continued during re-covery periods of67 to 110 minutes in duration. During both sets ofexperiments serialmeasurements ofthe

acid-base factors of blood and of the concentration of

(3)

J. R. ELKINTON, R. B. SINGER, E. S. BARKER, AND J. K. CLARK

trolytes in plasma were made on samples of arterial or

cutaneous arterialized blood (15, 16). In a few

in-stances ions other than bicarbonate were measured in venous serum. Simultaneously measurements were made

of the renal clearance of electrolytes and of renal

hemo-dynamics, to be described in a subsequent paper (4).

Heparinized blood for acid-base studies was collected

and handled with precautions to prevent loss of CO2 (10, 16). One portion of each sample was centrifuged

anaerobically, and the plasma drawn off and stored in a

greased syringe for analysis of total CO2 by means of

the manometric Van Slyke and Sendroy apparatus (17), sodium and potassium with a Barclay internal standard

flame photometer (18), and chloride by a titrimetric sil-ver iodate method (19). The remaining portion of each

sample of blood was stored in the capped syringe in an

ice bath until aliquots were taken for determination of

hemoglobin (20), and of hematocrit value, pH and whole blood CO2 by a modification of the Shock and Hastings

microtechnique (21, 16). In this modification visual colorimetric comparison of pH at 370 C. was replaced by

reading in the Klett-Summerson photometer, with a green filter, in a manner similar to that described by

Van Slyke, Weisiger, and Van Slyke (22). The

stand-ard error of this measurement was 0.009 pH unit for a single reading; these measurements, however, were

always made in triplicate. Venous blood, when used,

was allowed to clot, was centrifuged under oil, and the serum was drawn off for analysis of sodium, potassium, and chloride. By use of the Singer-Hastings nomogram

(15) and the Henderson-Hasselbalch equation,5 values werederived (from the observed values of plasma pH as

measured in whole blood and from CO2 content of plasma) for three additional values: arterial CO2 pres-sure, plasma bicarbonate concenitrationi, and concentration of buffer base. Buffer base represents the cation equiva-lent of the sumof bicarbonate anid all other bufferanions.

CALCULATIONS

Generalprintciples. Transfers of sodium andpotassium

oftenhave been estimatedinrespect tocalculated changes

in "extracellular" fluid as equated with the chloride space (1, 24). In essentially the same manner it is

possible to calculate from the changes in total

"extracel-lular" bicarbonate the transfers of hydrogen ion into and outof the chloride space, when allowance has been made for the uptake or release of hydrogen ion by the

non-bicarbonate buffers in red cells and plasma, and for the excretion of hydrogen by the kidney. The method of calculation employed here is similar to that described

From theHendersoni-Hasselbalch equation:

pH = _pKj' +_log

(HC03

)

where

pK,' = 6.10at370 C. (23)

f = 0.0314 mM perliterper mm. Pco2at 370 C. in

plasmaorsermmwith an averagewater content of 938g. perliter (23).

previously (10), but allowance is also made in the present report for shifts of chloride between red cells and plasma. This is necessary since under these

experi-mental circumstances such shifts maymake a significant

difference in calculated fluid volumes. These calcula-tions require values for initial and final "extracellular" fluid volumes. Such an initial volume was assumed and

the final volume was calculated by correction according to changein serum concentration and external balance of

chloride. This is probably at least as accurate for our purpose as direct measurement by substances such as

inulin or radio-sulfate since the changes in this chloride spaceand in the associated cationic transfersarerelatively

little affected by variations in the initial absolute "ex-tracellular" volume. The validity of the use ofthe

chlo-ride space is taken up under Discussion.

Transfers of hydrogen ion into and out of the chloride space, the red cells being included with this

"extracellu-lar fluid," may be calculated from changes in the total

amountof reacting buffer anions. Hydrogen ioncan

com-bine with bicarbonate or non-bicarbonate buffers (Buf-),

chiefly hemoglobin andplasma protein anions in blood, in

the following manner:

H+ + HCO3- = H2CO3 = H20+ CO2

H++Buf- HBuf

(1)

(2)

The CO2 is volatile, HBuf is non-volatile. Inmetabolic

acidosis the influx of strong acid provides hydrogen ion

which must be buffered by decrease of both bicarbonate

and other buffer anions as shown in Equations 1 and 2.

The excess H+ which reacts with dissociated buffer anions, Buf, produces an equivalent rise in the

undis-sociated acid, HBuf, and this excess H+ can be identified as remaining inthe chloride space in the form of a

non-ionized acid. On the other hand, the excess H+ reacting with HCO3- ordinarily cannot be so identified, since the

non-metabolic carbonic acid formed is broken down into

water and carbon dioxide which is very promptly

elimi-nated through the lungs. The excess H+ therefore is accountable as a measured or calculated decrease in the sum of the buffer anions, or in their cationic equivalent, the buffer base, BB+. In primary respiratory disturb-ancestherespective changesinHCO,-and Buf-arein

op-posite directions, instead of in the same direction. When

whole blood is saturated i1t vitro with air-CO2 mixtures each increase or decrease in HCO3- is balanced by an

equal and opposite change in Buf-. The reactions

repre-sented in Equations 1 and 2 are reciprocal reactions, and the buffer base content of whole blood is constant,

al-thoughthe chloride shift resultsinalimited redistribution of the buffer base between plasma and red cells.

The situation is more complicated when such changes

are considered in vivo, with at least four phases to be

accounted for simultaneously. These include red cells, plasma, interstitial fluid, and "intracellular" fluid. In addition, effects of renal excretion must be considered. By means of appropriate measurements all of these

fac-tors can be calculated in such a wayas to yield a

(4)

This value represents hydrogen ion that must be

ac-counted for in respect to a change in the total quantity

of "extracellular" plus red cell buffer base,

ABBe,+,

cor-rected for the externalnon-respiratory balance of hydro-genion, bH+. Itis calculated as follows:

AH+ = ABBer+-b+. (3)

Although this is designated as hydrogen moving in one direction, it may also represent bicarbonate moving in the opposite direction across the phase boundary; the two cannotbe differentiated. IfAH,+ispositive, one may refer to a transfer of hydrogen ion into the cells, and if negative, out of the cells. It should be understood,

however, that these opposite changes may represent only a change in the rate of movement of the ion in a con-stant direction rather than reversal of its movement. The continuing metabolism present in the dynamic

equi-librium which we designate as a "steady state" provides

acid products which cause a steady movement of

hydro-gen ions out of the intracellular fluid. In response to

respiratory acidosis one may calculate a positive AH,+,

and describe it as a shift of hydrogen ion into cells. It

may represent, however, a decrease in the steady rate at which the ion is moving out of cells, a process that would have an identical effect on the intracellular and

extracellular fluid.

The method of calculation of ABBer+ was similar to that described previously (10), with certain modifications oc-casioned by the fact that the chloride shift cannot be

neglected in respect to these ionic transfers. The al-tered quantity of buffer base was derived from

ABBer+ =

AHCO3er-

+

ABufb-,

(4)

where

ABufb-

is the change in non-bicarbonate blood

buffer anions as calculated below in Equation 9, and

AHCO3er-

is the change in total "extracellular" plus red cell bicarbonate calculated as follows:

AHCO3e

- =

AHCO3

- +

AHCO3

- +

AHCO32.

(5)

For plasma, interstitial fluid and red cells, indicated by subscripts p, f and r, respectively, the change in the amount of bicarbonate between observation 1 and 2 is given by the equation:

AHCOi-

=

_V2(HCOi-)2

-

Vl(HCO-)1,

(6) where V1 and V2 represent initial and final volumes,

re-spectively. The bicarbonate concentration in plasma,

(HCO3-)

, was calculated from the observed plasma

total CO2 concentration and pH, by means of the

Hen-derson-Hasselbalch equation. Redcell "bicarbonate" (in-cluding carbamino CO2) was calculated from the whole blood total

C02,

plasma total CO2 and red cell volume,

the appropriate subtraction being made for carbonic acid on the basis of the solubility coefficient and CO2

pressure. The whole blood CO2 concentration was cal-culated from the plasma CO2 concentration, pH, and whole blood hemoglobin concentration by means of the line chart of Van Slyke and Sendroy (17), on the

as-sumption of complete oxygen saturation in the arterial

orcutaneous blood. In the experiments where cutaneous

blood samples were used the plasma CO2 concentration

was calculated from whole blood CO2. Interstitial fluid bicarbonate concentration, (HCO-)f, was derived from

(HCOO-),

by the following equation:

(HCO3-)f

= 1.16

(HCO3-)p.

(7) The factor 1.16 is based on a Donnan ratio of arterial plasma to interstitial fluid bicarbonate concentration, ex-pressed per kilogram of water, of 0.91, and a water con-tentof935 grams per liter in plasma and990 grams per liter in interstitial fluid (25). The rather low value of this Donnan ratio as compared with the usually quoted one of 0.95 or 0.96 is explained by the relatively large arterio-venous difference for bicarbonate. The corre-sponding factors which were used for the concentrations of Cl, Na and K were as follows: 1.08, 1.015, and 0.97, respectively (26,27).

Initial control values of blood volume, Vb, and extra-cellular fluid volume,

V,,

were assumed, 80 ml. per Kg.

body weightfor the former (28, 29) and 200 ml.per Kg.

body weight for the latter (10). Successive values of

blood volume were calculated from hemoglobin

concen-trations, (Hb), by the equation:

Vb2 Vbb (Hb)<

(8)

For each bloodvolume the corresponding plasma and red cell volumes,

Vp

and Vr, were derived with the aid of the observed hematocrit value. The initial interstitial fluid volume was obtained as the difference between the initial assumed extracellular and plasma volumes.

Sub-sequent values of interstitial fluid volume, Vf, were calculated in terms of the chloride content of blood and interstitial fluid, allowance being made for loss of chlo-ride in the urine and shifts between blood and inter-stitial fluid; the assumption was made thatthere was an

equilibrium in chloride concentration between blood and interstitial fluid at the time of sampling. Where whole blood chloride was not measured an initial value was calculated and transfers between plasma and red cells were taken in accordance with the pH changes and the average transfer per unit change in pH observed in the arterial blood experiments. For each observation after the initial one it was thus possible to calculate (a) the amountof chloride in interstitial fluid, on the assumption that no exchange occurswith tissue cells, (b) the inter-stitialfluidchloride concentration, (Cl-)f,as 1.08 X (Cl-) , and (c) the interstitial fluid volume as the quotient of amount divided by concentration. These volumes were used to calculate not only the bicarbonate distribution,

but also the distribution of extracellular sodium and

potassium.

The final term in Equation 4, the change in non-bi-carbonate blood buffer anions, was calculated from the observed pH changes, hemoglobin concentration, and

as-sumed blood volume asfollows:

ABufb = [7.0 Vp +2.3 (Hb)bVb] ApH. (9)

(5)

17J.

R. ELKINTON, R. B. SINGER, E. S. BARKER, AND J. K. CLARK

onbuffer data for normal human blood of Dill, Edwards, and Consolazio (23), as adapted in a slightly different form (15).

With the value for ABBer+ thus obtained, the

non-re-spiratory hydrogen ion balance, bH+, mustbe calculated in

order to solve Equation 3 for change in intracellular

hy-drogen ion. In the absence of intake this hydrogen ion

balance is calculated from the urinary excretion of am-monia and titratable acid, UVNH4+ and UVTA, which measure hydrogen excreted through the kidney, and of

bicarbonate,

UVHCO03,

which is equivalent tohydrogen ion

added to the body:

be

= - [(UVNH4+ +UVTA) -UVHCO3<]. (10)

Hyperventilotion

Mean changein:

+0.20

Plasma

pH

o

pH ~~-0. 1oJ

mean

.S.. P.

Given the values for ABBer+ and for bHa thechange in intracellular hydrogen ion, AH,+, is then calculated from Equation 3.

RESULTS

I. Effects on blood and plasma

All

results have been evaluated by standard

statistical procedures. These results

are

presented

in Tables

I

A and

I B

and in

Figures 1 and 2.

Acid-base factors of blood and

plasma

(Figure

1).

In response to

voluntary hyperventilation the

CO2inhalation

+0.24 +0.03 -0.08 +0.01

*0.019 *0.013 *0.014 *0.020

<.OPI .07 .002 >.58

Total C.02,.

[CO2]p

.

M./ 1. -10

meon S.S.

p-Whole blood

+21

Buffer

base,

o-A

_[BB]b

_mean

mEq./1- S...

p--6.9 -0.2

* 0.74 + 0.34

<.001 .58

-1*

0 +I

*0.7 *0.5

> .58 .12

+ 1.6 -1,1

* 0.35 *0.42

.006 .05

_ I _ I

*0.7 +0.8

.19 .25

Plasma

CO2 pressure,

A_Pco2 mm.Hg

-24 -3

i 1.5 *1.I

(.001 .03

+15 -6

* 1.5 *2.2 .ool0 .04

before 0 after FIG. 1. ACUTE RESPIRATORY ALKALOSIS AND ACIDOSIS: MEAN CHANGES IN PLASMA PH, PLASMA CO2 CONTENT, WHOLE BLOOD BUFFER BASE

CON-CENTRATION, AND CO2 PRESSUREIN PLASMA

The mean for each group of changes from the individual mean control values are plotted for the end of the periodof stimulus and the end of the recovery period. The values for the mean, its standard error, and the

probability that the mean change is significantly different from the

con-trol, are given below the curves. The mean changes that are statistically significant are represented by open circles.

+101

o

2

-20.

kJ

Time

meon

S.C. P.

L

(6)

Mean

change

in

plasma

conc:

of

Potassium

+0.3

ACK]

p _

nEq./L. -03 mean

*.e.

p.

Sodium

m[NE]p

mEq./1. -2

mean -1.0 +0.3

s.e. *0.54 £0.72

p. .13 >.58

Hyperventilation

CO2 inhalation

Chloride

Ajc1 I_p

mEq./I.

Ti

me:

+2

-2

s.e.

p-+1.6 -0.3

*0.54 *0.85

.04 >.58

L

before 0 after

-0.4 0

*0.38 *0.56

.32 >Z8

before0 after

FIG. 2. AcuTE RESPIRATORY ALKALOSIS ANDAcIDosIs: MEAN CHANGES IN THE PLASMA CONCENTRATION OF POTASSIUM, SODIUM, AND CHLORIDE

The data are presented as in Figure 1.

pH

rose

(mean

= ₊

0.24 pH unit), the CO2

con-tent

fell

(- 6.9

mM

per

L.), the

pressure

of CO2,

Pco2,

fell

(-

24 mm.

Hg),

and the buffer base

concentration,

(BB+)

b, was constant.

These

changes define

a

primary respiratory alkalosis

or

carbonic

acid

deficit.

After the stimulus these

values

returned

to or

towards the control levels.

In CO2

inhalation the

pH fell (- 0.08 pH unit),

the

CO2

content rose

(+ 1.6 mM

per

L.),

the

Pco2 rose

₍₊

11 mm.

Hg),

and the buffer base

concentration

was

essentially unchanged

(-

1 mEq.

per

L. These

findings

are

characteristic of

a

primary respiratory acidosis

or

carbonic acid

excess.

After the inhalation, elevated

values for

Pco2

and total CO2 returned to

normal, actually

overshooting

to

reach

a

significantly

decreased

value

at

the end

of the experiments.

Since

the

mean

changes

in

pH and Pco2

were greater

dur-ing voluntary hyperventilation

than during

CO2

inhalation, it is

apparent

that of the

two

proce-dures

the

former

was a

considerably

more severe

disturbance and

hence, in

comparison

to

CO2

in-halation, evoked

more extreme as

well

as more

consistent

responses

in

the body.

Plasma

concentration of other

electrolytes (Fig-ure

2). Slight changes

were

obtained in the

mean

concentrations

of

sodium, potassium,

and

chloride.

Although in

most

instances these changes

were

not

statistically significant, they

were

in

opposite

directions with the

two

stimuli and in the direction

predictable

on

theoretical grounds.

When

con-verted

from concentration

changes

to

changes

in

total extracellular

amounts many

of these

changes

are

significant.

The decrease

in

potassium

con-centration of

-

0.3 mEq.

per

L.

atthe

end of

hy-perventilation (significant

at

the

6 per cent level

but

not at

the 5

per cent

level)

is

in accord with

other observations in

experimental respiratory

alkalosis

(30, 31).

The

significant

increase

in

chloride

concentration,

+ 1.6

mEq.

per

L.,

oc-curring

in

respiratory alkalosis,

is

predicted

on

theoretical

grounds

from

the

effect of the

pH

rise

in

decreasing

the Donnan

ratio of red cell

chlo-ride

to

plasma

chloride

(23).

II. Calculated

effects

on ionic

transfers

in

the

total

body

fluids

These results

are

presented

in Tables II

A,

II

B,

III

A,

and

III

B,

and in

Figures

3 to

6,

inclusive.

In Tables

II A and II Bare

presented

the observed

balance

data,

the calculated ionic concentrationsin

-0.3 -0.3 +0.1 -0.1

*0a12 £0.15 £0.06 *0.89

.06 .10 .16 >.58

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+0.5

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(7)

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(9)

1-J. R. ELKINTON, R. B. SINGER, E. S. BARKER, AND J. K. CLARK

interstitial fluid, and the calculated volumes of the

several fluid phases, all of which are necessary to

the

calculation of

the ionic

transfers

shown in

Tables III A and III B.

Extracellular bicarbonate and buffer base, and

intracellular

hydrogen

(Tables III A and III B,

Figures 3 and 4). In respiratory alkalosis, at the

end of

the

stimulus,

the

total extracellular

bicarbo-nate changed by a mean of

-

136 mEq.

with

a

re-ciprocal change

in total

non-bicarbonate

buffer

anion of + 39 mEq.

In

respiratory

acidosis

the

same

changes

were + 32 mEq. and

-

12 mEq.,

respectively. The algebraic sum

of

these two

val-ues,

the change

in

total extracellular buffer

base,

was

-97 mEq.

in

respiratory

alkalosis and + 20

mEq.

in

respiratory acidosis.

All values except

the

last

were

significant

at

the 5 per

cent

level.

However,

from

the

end of

the

CO2 inhalation

to

the

end of

the

recovery

period

a

change

in

the

op-posite direction occurred

with the loss of 36

mEq.

Hyperventilation

10ta1 +50

8extracellulor"

bicarbonate

°

-, AHCO; -50

E j

-100

II

0)

CP 0

0

E

Cz

0

a,

Totol blood

non-bicorb.

buffer

ABuf_b

mean

$.S.

p

of extracellular buffer base. This recovery change

was

highly significant (p

<

0.001), a finding

add-ing

strong support to the contention that the

ini-tial response

during CO2

inhalation was real.

When these values are corrected for the small

amounts of

buffer base excreted with bicarbonate

and conserved by the secretion of acid and

am-monium by the kidney, they provide a

quantita-tive estimate of the transfer between the

extracel-lular and intracelextracel-lular phases of hydrogen ion in

one

direction or bicarbonate in the other (Figure

4).

Thus,

taken as change in the former, the

mean

change in intracellular hydrogen was

-

92 mEq. in respiratory

alkalosis

and

+ 22 mEq. in

respiratory acidosis. Once again, while the mean

change in intracellular hydrogen between control

periods and the end of the stimulus was

significant

in

the alkalosis group only (p =

0.005),

the mean

reciprocal change

from the

end

of

the stimulus

to

CO2

inholotion

C....

116

+39 +6. -12 I

i 5.6 *2.7 * 1.9 *2.6

.002 .093 .002 >56

Totol

"extracellulor"

buffer bose

aBe.

-100I

mean

CSe.

p

-97 -6 +20 -;6

*16.1 *11.3 *10.3 *10.3

.004 >.58 .090 .170

before 0 after before 0 ofter

FIG. 3. AcuTE RESPIRATORY ALKALOSIS AND Acmosis: MEAN CHANGES

IN TOTAL "EXTRACELLULAR BICARBONATE," TOTAL BLOOD NON-BICARBONATE BUFFER ANION,AND TOTAL "EXTRACELLULAR" BUFFER BASE

The dataare presentedasin Figure 1.

mean -136 -13 +32 -17

s.e. *19.8 *9.0 9.2 *6.9

p .003 .234 .017 .1

+50I4_

Time

1678

T&4 I

I

(10)

Hyperventilation

+5

mean +48 +11 -16 +4 s.e. ±15.4 ±14.0 ± a5 ±9.5

p .036 .523 .116 >.58

07

033

-3.0 -3.1

*0.87 *1.62

.019 .130

L ,

before 0 after before 0 after

FIG. 4. AcuTE RESPIRATORY ALKALOSIS ANDACIDOSIS: MEAN CHANGES IN

INTRACELLULAR HYDROGEN, SODIUM, AND POTASSIUM

The dataare presentedas in Figure 1.

002 inhalation ",:

the end of

recovery

following CO2

inhalation

was

highly significant (- 37 mEq.,

p

< 0.001).

Intracellular sodium

and

potassium

(Tables

III

A and III B, Figure 4). In respiratory

alka-losis sodium decreased

in

the chloride

space to a

far

greater extent

than

it

was

lost

in

the urine;

in

respiratory

acidosis sodium

increased

or was

un-changed

in

this phase.

On the assumption that

the chloride

space

is

essentially equivalent

to

the

extracellular

fluid,

these data indicate that sodium

moved into the intracellular phase by

amean

value

of + 48

mEq. during respiratory alkalosis;

dur-ing respiratory acidosis the

mean

change of

-

16 mEq.

was

in the opposite

direction

but

was not

statistically significant.

During

the

recovery

pe-riod in

each

type

of

disturbance

the intracellular

sodium returned

to

the control

levels; the change

from the end of the CO2

inhalation

to

the end of

the

recovery

period

being

+ 20

mEq.,

p=

0.04. Intracellular

potassium

exhibited

changes

that

were smaller in

magnitude,

the mean

changes

be-ing

-

3.8 and

-

3.0 mEq.

during respiratory

al-kalosis and

acidosis, respectively.

Only

the latter

value

was

significant, although

a

significant

de-crease

in

intracellular potassium

was

found

during

the

recovery

period in

respiratory

alkalosis.

The quantity

AHi

-

ANai

represents

the

sum

of

the

transfer

of

hydrogen

ion in

one

direction

and transfer of sodium

in

the opposite direction.

Sincepotassiumtransferswere small,AH1 ANaj

in

these experiments

was

essentially the

same as

the total cation

exchange.

The value for this

change

in

the

case

of

respiratory

alkalosis

was

140 mEq.

from the control

state to

the end of

hy-perventilation,

and 123

mEq. between

the end of

the

stimulus and the end of

recovery.

In the

case

of

respiratory

acidosis the

changes

in

this

quan-tity

were

37 and 56

mEq., respectively.

All

four

of these

figures

are

highly significant (p < 0.001)

and indicate that the calculated

exchange

of

hy-drogen

for

sodium

across

the

phase boundary

un-der conditions of these

experiments

was not a

random

phenomenon.

Interrelationships of cation transfers

are

illus-trated

in

Figures

5 A and 5

B,

in which the

pat-tern

of

response in

each

group of

experiments

is

shown.

In

respiratory

alkalosis

(Figure

5

A),

the

change

in

total

extracellular bicarbonate

(de-+501

O.

-SO

-100

mean

s.e.

p

C 0

00

0

a

=

0-4._

._

c

0 cm

E

a C

0

4,

Hydrogen

AHi

Sodium

ANot

Potossium

A_K1

Time

-92 -4 +22 -15

*16.4 *10.7 *10.3 * 9.0

.005 >.58 .090 .150

mean -3.8 -9.8 s.e. * 2.40 *3.C

p .185 .C

(11)

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(13)

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(15)

picted on the left in each figure) was mostly due to

extra-renal, i.e., respiratory, loss from the body;

a

relatively

small

portion was excreted by the

kid-ney in

association

with

the fixed

cations,

potassium

and sodium.

On

the

right

in

each

figure are

shown

the

concomitant transfers

of

cations.

The

large respiratory losses of bicarbonate as carbonic

acid

required

hydrogen

ion

which came in part

from the

protein buffers of the blood and in part

from the body cells.

The

decrement of cellular

hydrogen

was replaced in part by an increment of

cellular

sodium;

some

portion

of

the remainder

was

associated with

a

transfer of

undetermined

ir +50

in VP c

a

C0

c L

._0 a

E

C,.

-50

-Ioo

-150

anion

(see below). Changes in cellular potassium

were

relatively insignificant.

In

respiratory acidosis (Figure 5 B), the

op-posite

pattern of

electrolyte transfers

obtained,

though to a somewhat lesser degree. The increase

in total extracellular bicarbonate resulted from the

CO2 inhalation rather than

changes

in renal

excre-tion. The

associated

hydrogen

ion was taken up

by the protein buffer anions of the blood and by

body cells.

Simultaneously cellular sodium

di-minished while cellular potassium was essentially

unchanged.

Thus our derived results indicate that acute

re-Hyperventilation

0 +30 +120

Total

bicarbonate

o +30 +120

Total

cations

FIG. 5A. RESPIRATORYALKALOSIS: THE MEAN SUMMATED PATTERNSOFRESPONSE

OF TOTAL EXTRACELLULAR BICARBONATE AND OF TOTAL BODY CATIONS, TO HYPER-VENTILATION

Changes are plotted cumulatively from the onset to the end of the stimulus and to

the end of the recovery period. On the left the dotted line indicates change in

bi-carbonate by renal excretion; the solid line indicates the total change in extra-cellularbicarbonate, thedifference being extra-renal orrespiratory. On the right are

depicted the corresponding changes in cations. The dotted line represents renal

ex-cretion of sodium and potassium, the dashed line respiratory loss of hydrogen from

bloodbuffer proteins, and the heavy line total hydrogen loss. The difference between the two latter represents transfer of hydrogen from total body cells. The associated changesin intracellular sodium plus potassium and in intracellular sodium alone are

indicated above by the solid line and dashed and dotted line, respectively.

Loss of extracellular bicarbonate by the respiratory route greatly exceeds that by

the renal. Associated with the loss ofbicarbonate through the lungs is the hydrogen ion drawn in part from theprotein buffers of the blood and in part from body cells. The latter decrement is partially replaced by transfer of sodium into cells.

Time

-min.:

(16)

CO2

inholatton

FaHAHCO

..fO3*0exr-e 3

renaly- N

0 +30

AH+

ANa1 +Kj

+120

Total

bicarbonate

0 +30

Total

cations

FIG. 5B. RESPIRATORY AcIDosIs: THE MEAN SUMMATED PATrERNS OF RESPONSE OF

TOTAL EXTRACELLULAR BICARBONATE AND OF TOTAL BODY CATIONS, TO CO2

IN-HALATION

Dataare presentedas in Figure 5A.

Extracellular bicarbonate is increased in respiratory retention, and hydrogen is taken

up by blood protein buffers and by body cells. Simultaneously the body cells lose

sodium.

+100'

+50

ANao

+

_AKi

0.

mEq.

-50

0

regr.

coef.

a -0.500, p(0.001

95 % C.L. a _{-0.365 to-0.635}

A.

A4 A Ab

A

o

N.

-100

-50

0

+50

+100

+150

AH,+

mEq.

FIG. 6. RESPIRATORY ALKALOSIS ANDACIDoSIS: RELATIONOF CALCULATED TRANSFERS

OF CELLULAR HYDROGEN TOTHOSE OF CELLULAR SODIUM PLUS POTASSIUM

Solid circles (representing alkalosis) and triangles (representing acidosis)

indi-catechanges duringtheperiodofstimulus; opencircles andtriangles indicate changes

from the end of the stimulus to the latter part of the recovery period.

Theinverse correlation between thetwo variables indicates a reciprocal relationship between transfers of these cellular cations. The relationship, as indicated by the

re-gression coefficient (solid line), is approximately that of two hydrogen ions in ea-change for onesodium orpotassium

iop.

+501

4

Vf c E

C

I 0

U

0

o

O-E 0

-50

(17)

spiratory

disturbances of acid-base equilibrium

in-volve transfers of hydrogen between the

extracel-lular and

intracellular phases of the body fluids.

As shown in Figure 6, these net transfers of

cel-lular hydrogen were reciprocally related to those

of cellular sodium plus potassium; of the

trans-fers of

these

two

latter cellular cations, those of

sodium predominated.

This inverse correlation

is

highly

significant (r

= -

0.868, p < 0.001).

The regression coefficients for these sodium and

potassium changes

on

the reciprocal hydrogen

change

were

statistically significant

in both sets

of

experiments

and were not

significantly

different

from

each other.

The regression coefficient for

the

pooled

values from both groups was

-

0.500 with

a

standard

error

of

+

0.0634, thus

indicating

a

95 per cent

chance that

the true value lies

be-tween

-

0.365 and

-

_0.635,

The

magnitude of

this

relationship

therefore suggests a two-for-one

exchange, i.e.,

under these particular conditions of

either acute

respiratory

acidosis or alkalosis

ap-proximately

two

hydrogen ions

move for one ion

of sodium

and potassium

considered together.

Transfers of undetermined anion

may explain the

maintenance of

electroneutrality

of

the

extracel-lular

fluid

in

the presence of

this

unequal exchange

of cations.

Calculated

changes

in undetermined

ion

approximated

closely the excess of hydrogen

transfer over the

opposite

transfer of sodium,

dur-ing stimulus and

recovery.

Since

essentially the

same

factors

comprise both calculations, the

unde-termined

anion data are

not

presented although

they

appear to represent

a

valid deduction

of one

link

in the

ionic transfers that

occur

in response

to

these

respiratory

acid-base

stimuli.

DISCUSSION

Validity of

the

calculations

The principal assumption underlying the

cal-culations is that

changes

in the volume of

extra-cellular fluid may be

quantitated

from

exchanges

of

chloride, i.e.,

the

chloride space is

equated with

the true

extracellular volume. We

recognize

that

chloride is present in

some

cells and

that, strictly

speaking,

the chloride space

cannot

be

equated

with the "true" extracellular space.

However,

there is

an

increasing body

of

evidence

to

indicate

that larger

molecules,

such

as

inulin,

fail

to

pene-trate

that

portion

of

the

extracellular

fluid which

consists of connective tissue or collagen (32,

33),

and that this fact may well explain some of

the

discrepancies between the distribution of

inulin

and of

the smaller ions, chloride, bicarbonate, and

sodium

(34-36, 10).

We

have discussed this

problem in detail in an earlier paper

(10).

In

the absence of a better measure of

the

volume of

the

"true" extracellular

fluid,

for purposes

of

cal-culation the chloride space is taken here as its

ap-proximate equivalent.

It

is also

recognized that

the

movement of ions out of the chloride

space

does

not

necessarily

mean

that

they

enter

cells

in

general

or

cells of any

particular tissue.

It is

not known to

what

extent transcellular

"pools"

such

as

cerebrospinal

fluid and

gastrointestinal

secretions

may

participate

in

these exchanges.

Sodium

has

been shown

to

be

readily

mobilized

from bone in

certain animals

(37, 38).

But

the

fact

that the cells of

the

bulk

tissues,

such

as

skele-tal

muscle, contain little chloride and much

protein

(a

potential buffer), and the fact

that

muscle

analyses by various

workers have

indicated

changes

in

ionic

concentration

in response

to

acid-base disturbances,

make

it

reasonable

from the

physiologic

standpoint to label as

"intracellular"

those

ions

which

move outside the

chloride

space.

Nevertheless,

it

is

pertinent

to

assess

the

effect

on our

derived data of

possible

shifts of

chloride

between the

true extracellular and intracellular

phases of the body fluids. Those between red cells

and extracellular fluid have been taken into

ac-count. If chloride did

move

into cells under either

of

our

experimental

conditions,

the calculated

in-crements in

cellular

sodium would be

too

small

or

decrements too

large;

conversely,

if

chloride

had

moved

out

of cells

the

reverse would be

true.

However, since the

calculated

transfers

of cellular

sodium

and

hydrogen

were

in

opposite

directions

in 19 out of 22 periods of stimulus and recovery,

diminution

or

enhancement of

change

in

cellular

sodium due to cellular chloride shift would be

as-sociated with the

opposite

effect

on

cellular

hydro-gen.

Therefore

the

occurrence

of such shifts of

chloride,

unallowed-for in the

calculations,

would

alter the

quantitative

reciprocal

relationship

be-tween

changes

in

cellular sodium and

hydrogen,

but would

not

invalidate

the evidence that

trans-fers of

one or

both cations took

place

across

the

phase

boundary.

The calculated difference between

change

in

(18)

tracellular buffer base and the external balance of

bicarbonate is

a measure

of

transfer of

hydrogen

ion in

one

direction in

respect to

cells

or

transfer

of

bicarbonate ion in the other direction. For

con-venience

we

have

chosen

to

label the

findings

as

movements

of

hydrogen ion.

While it

seems

somewhat

more

probable

tous

that this

represents

the

transfer

involved, it is recognized that the

dis-tinction

cannot

be made by

present

methods and

that the

net

result

would be

the

same

if

instead

bi-carbonate ion

moved in the opposite direction.

The

major role of intracellular ionic transfers

in

buffering extracellular pH changes in acid-base

disturbances.

The results of these experiments

indicate

very

strongly that the cells of body tissue

provide

a

major buffer

system

against the effects

of

acute

acid-base disturbances of the respiratory

type.

Ninety

per cent or more

of the

change in

effective extracellular bicarbonate

was

extra-renal

and

was

changes in hydrogen uptake

or

release by buffer protein in the blood and by

ionic transfer into,

or out

of, cellular fluid in the

body;

only

a very

small

fraction

was

accounted

for

by acid-base adjustments in the kidney

over

the

short

periods

of

time

of these

experiments.

Exchanges

of hydrogen ion with blood buffer

sys-tems

and

in

the

kidney have heretofore received

much

attention;

exchanges of hydrogen ion with

intracellular buffer

systems

have

not

been

con-sidered

so

extensively.

Earlier workers had demonstrated

experimen-tally

that

a

major

portion

of

CO2 withdrawn from

the

body by hyperventilation,

or

retained in

the

body

by inhalation,

came

from,

or was

added

to,

tissues

other

than blood

(39-41 ). On the

assump-tion

of

an

unchanged

basal

R. Q., Rosenbaum

(42) calculated that during hyperventilation 21

to

47

percent

of

expired

non-metabolic

CO2

came

from

outside

the chloride-corrected thiocyanate

space;

conversely, during CO2 inhalation, 12

to

56

per cent

of

the CO2

was

retained in

this

space.

These

studies

clearly

indicated

that

the

intracellu-lar

fluid shared

in

the

buffering of

respiratory

acid-base

disturbances)

but

there

was not

sufficient

ap-preciation of the obligatory

nature

of associated

transfers

of

hydrogen

and other

cations.

These associated

ionic

transfers

are

delineated

in

our

experiments and

in

the

important

paper

by Giebisch, Berger, and Pitts (9) which has been

published

since

our

experiments

were