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JOURNAL OF NUCLEAR MEDICINE 7:827-836, 1966
Muscle Electrolyte Composition Determined by
Neutron Activation
A Preliminary Study of Dehydration in Infants.'
J. Dubois,2 J. Colard,3 and H. L. Vis
Brussels, Belgium
INTRODUCTION Because muscle tissue is abundant, easily approachable and of relatively constant composition, both hydrosaline and energetic metabolism can be easily studied in the muscle area. The activation analysis method, recently applied by Bergstrom (1) to the study of this material allows electrolytes (Na, K, Cl) and phosphor determina tions, on small muscular fragments taken by needle biopsies. This method has been adapted to the irradiation characteristics of the BR1 and BR2 reactors of the C.E.N. Mol (Belgium). In order to increase the speed and precision of the determination of induced activity, a new technique of measurements was devised. The purpose of this work, of which preliminary results are described in this paper, is a direct observation of the muscular hydrosaline equilibrium in some states of infantile dehydration.
MATERIAL AND METHOD
Needel biopsy technique. The technique of the needle muscular biopsy was
described by Bergstrom (1), who modified the Pollen and Bickel needle, which is pointed and hollow with a window at its distal end. A cylinder with a sharp rim fits into the needle and permits the sampling of material engaged in the window. Finally, a stylet may be introduced in the cylinder in order to extract the sample (Fig. I).
‘This work was done under contract Euratom-Université de Bruxelles-Université de Pise 026-63-4 Biac. 2Service et Laboratoire de Pédiatrie (Prof. R. DUBOIS) Université Libre de Bruxelles (Belgium). 3Centre d'Etude de l'Energie Nucléaire(Mol)—Belgium. 827
828 DUBOIS,COLARD,VIS
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II 4
10
12
13 Fig. la. Needle1 used for muscle biopsy similar to that used by Bergstrom (1) but modified for pediatric investigation.
‘Manufactured in Belgium b@ SIMAL (CEMBLOUX).
830 DUBOIS, COLARD, VLS
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Fig. lb. From right to left: 1) Hollow needle window diameter (0.6 cm). 2) Cylinder with a sharp rim (diameter 0.2 cm). 3) Stylet (diameter 0.15 cm).
Half lifeCapture
cross section act.)-yf3Na2415.0 (oP.
h0.52 barn1. 100%1.39\lev100%K4212.5 2.75MevMev100%
h1.17 (^) barn1.53Mev18%3. 18%Cl'8 1.98\levMev82%
p3237.
mm.
143 days0.
barn
0.21 barn1.
2.15Mev\Iev
—31%
1.71Mev
Mev Mev @\Iev31%
MUSCLE ELECTROLYTE COMPOSITION AND NEUTRON ACTIVATION 831
The isotopes produced are Cl38 with a half-life of 37.5 mm. emitting 1. Mev and 2.15 Mev gamma rays and Na24 with a half-life of 15 h. emitting 1. and 2.75 Mev gamma rays (Table I). A sample of 1 ml of a solution of NaCl of known concentration, (O.OIM), is irradiated simultaneously with each group of biopsies. After irradiation, this
solution is diluted to 10 ml and two samples of 0.5 ml are pipetted as standards.
The amounts of Na and Cl in the biopsies are determined by comparison with
these standards.
In order to differentiate the gamma ray characteristics of Na24 from Cl38,
the measurements are made by gamma spectrometry.
The equipment used requires a well-type Nal crystal detector of 3―x 3―.
The gamma spectrum is accumulated in the memory of a 400 channel analyser
(Intertechnique) for five minutes and automatically recorded on a magnetic tape. The following sample is measured and recorded immediately afterwards. A perfect stability of the spectrum is obtained by means of a drift stabilizer, “Stabimat―,centered on the Na24 2.75 MeV photopeak.
The quantitative measure of Na24 is calculated from the 2.75 MeV photopeak
rather than the 1.37 MeV photopeak, which is distorted by the K42 1.53 MeV
photopeak.
A reference spectrum of Na24 is recorded on a magnetic tape in the same conditions as those of the samples. To obtain the relative activities, this spectrum
or one of its multiples is subtracted, by means of an intermediate computer (RG
TABLE I
ISOTOPE OF SoDIuM, POTASSIUM, CHLORINE AND PHOSPHORUS OBTAINED BY NEUTRON ACTIVATION.
MUSCLE ELECTROLYTE COMPOSITION AND NEUTRON ACTIVATION 833
After determination of Na and Cl, the samples and two standards of KHOPO
are irradiated for ten minutes with a BR2 flux of 10'@neutrons/cm2/sec.
Afterwards, they are kept for 24 hours to obtain a sufficient decrease of Cl
and Mn5°activity. The biopsy is then digested in 3 ml of 7N warm nitric acid
containing a carrier of KH2PO4 102M. Three samplings of the solution are pipetted off on counting trays and dried under an infrared lamp.
The counting trays are then placed under an antracene crystal mounted on a
photomultiplier connected to a monochannel analysor.
To measure K42 (max f.@emission : 3.5 MeV ), and in order to avoid the de
tection of P32 ( max. : 1.7 MeV ) the monochannel analysor window width is 2
MeV and the lower level is 2 MeV. However, the result must be corrected be
cause of the presence of sodium-24. In reality, the gamma rays of 2.75 MeV
can be detected by the Compton effect in anthracene. Moreover, the simultaneous
detection of Na24 $ ray (1.39 MeV) and of the Compton event of one of the y radiations of Na24 can give an impulse exceeding the 2 MeV threshold fixed at the monochannel. In the material studied, this error can reach 25% of the total activity. The measurement of @32is done on the same counting trays after 10 days, allowing time for the decay of K42 and Na24. In this case, the monochannel window set from 0,4 to 1,8 MeV does not de tect the S35 and Ca45. The K and P contents of the muscular sample are calculated in reference to the standards.
PRELIMINARY RESULTS AND DISCUSSION Preliminary results are shown in Table II and III. Table II points out the values obtained by chemical and neutron activation methods in normal children,
and shows that there is no significant difference between the results obtained
by the two methods. We emphasize the fact that all our biopsies were done on children, because the values are different from those obtained in adults and cannot be compared. Similarly, Bergstrom's values for normal adults by the neutron activation method are identical with those found by other authors (3,4,5,6,7,8) by chemical
methods and we may therefore assume that the values found by the two methods
are comparable. Table III shows quantities of total water and electrolytes found in children with two different acute dehydration states (Isotonic and hypertonic dehydra tions). We have classified patients as hypertonic if serum sodium concentrations are 150 mEq/liter or higher. The preliminary results seem to indicate a rapid participation by muscular tissue in water and electrolyte movements which are characteristics of the above states. Moreover, these variations are generally similar to those obtained in the plasma. This seems to show that the muscular water osmolarity is well depicted by the plasmatic ionogram. In certain conditions, however, the interpretation of the data is ambiguous.
834 DUBOIS,COLARD,VIS
Our results are expressed, as usual, in relation to dry fat free solid. Thus
we assume that the dry tissue remains constant during all pathological processes. This assumption is probably true in acute states of dehydration occuring in pre viously healthy children, but we have noted that this material was modified in some cases of chronic malnutrition (7). In these last cases, the extracellular protein material composition is quan titatively different from that of normal muscle; the collagenous fractions become greatly augmented in comparison with the other extracellular protein fractions. Moreover the relation of total extracellular proteins to intracellular proteins changes, that is, referring the amounts of electrolytes to dry fat free weight will lead to a false interpretation. For example, the potassium values will be over estimated. We have to admit, therefore, that reference to fat-free solids will be inade quate in some circumstances. As do Lilienthal and his colleagues (9), some authors suggest expressing the results in terms of non-collagen nitrogen, that is, in terms of intracellular pro teins and we have indeed demonstrated that non-collagen nitrogen remains con stant even in severe pathological cases where collagen nitrogen is modified (10). It is impossible, however, to determine non-collagen nitrogen accurately on small fragments taken by needle biopsy. A surgical biopsy is necessary, but cannot be carried out in most cases.
TABLE II
i\ltSCt'LAR COMPOSITION IN TOTAL WATER, ELECTROLYTES AND PHOSPHORUS IN NORMAL CHILDREN, MEAN AND S.D.' Column A: results obtained by neutron activation. Column B and C: results obtained in two groups of children by chemical methods (7).
A B C
Total water 354.7 ± 6.2 342.4 ± 5.0 346.2 ± 5. —9 cases Na 18.4± 1.7 25.8± 1.0 16.8± 1. — 10 cases
K 44.4± 1.2 42.2± 1.0 44.7± 1.
—9cases (1 13.5± 1.5 15.4± 1.1 17.4± 0. — 10 cases 26.6± 1.4 29.9± 1. —8 cases —6 cases —10 cases
‘Theresults are expressed in g. for water and in mM for electrolytes and phosphorus, per 100 g. dry fat free solid.
836 DUBOIS,COLARD,VLS
SUMMARY AND CONCLUSIONS Following the work of Bergstrom in the adult, we have described a method of analysis of muscular electrolytes and phosphorus in children. By comparing our results with those obtained by the chemical technique we came to the con clusion that this method is reliable. An analysis of needle biopsy specimens of muscle from children with differ ent degrees of dehydration suggests that the muscular tissue is rapidly involved in acute hydro-electrolytic disorders. The preliminary data will be checked by the study of a larger series of cases. Although the use of such a method is limited, it remains the only one which allows a direct study of tissue in pediatric pathology.
REFERENCES
- BERGSTRoM, J.: Muscle electrolytes in man. Scand. J. Gun. Lab. Invest. 14, suppl. 68,
- FLEAR, C. C. T. AND FLORENCE, I.: A rapid and micro method for the analyses of skeletal muscle for water, sodium, potassium, chloride and fat. Gun. Chim. Acta. 6:129, 1961.
- BARNES, B. A., GoRDoN, E. B. AND COPE, 0.: Skeletal muscle analysis in health and in certain metabolic disorders. The method of analysis and the values in normal muscle. J. Clin. Invest., 36:1239, 1957.
- DICKERSON, J. W. T. AND WIDDOWSON, E. M.: Chemical changes in skeletal muscle during development. Biochem. 1., 74:247, 1960. 5. LITCH.FIELD,J. A. AND GA.r'Dr.E,R.: The measurement of the phase distribution of water and electrolytes in skeletal muscle by the analysis of small samples. Gun. Sci., 17:483,
- MULDOWNEY, F. P. AND WILLIAMS, R. T.: Clinical disturbances in serum sodium and potassium on relation to alteration in total exchangeable sodium, exchangeable potassium and total body water. The value of muscle biopsy analysis in diagnostic and management. Am. J. Med., 35:768, 1963.
- Vis, H. L.: Aspects et mécanismes des hyperaminoaciduries de l'enfance. Arscia Bruxelles et Maloine-Paris Ed., 1963.
- WILSON, A. 0.: Electrolyte content of muscle samples obtained at surgical operations. Brit. I. Surg., 43:71, 1955.
- LILLIENTHAL, J. L., ZIERLER, K. L., FOLK, B. P., BUKA, R., AND RILEY, M. J.: A refer ence base and system for analysis of muscle constituents. I. Riot. Chem., 182:501, 1950.
- DuBoIs, J., COLARD, J. et Vis, H. L.: Etude des méthodes de dosage des electrolytes, du phosphore, du materiel azotéet des acides organiques musculaires. Emploi de l'activation neutronique. (to be published).
