Life at the Cell and Below-Cell Level. The Hidden History of a Fundamental Revolution in Biology
"Dr. Ling is one of the most inventive biochemist I have ever met." |
Chapter 7. Legacy of the Nearly-Forgotten Pioneers (p. 35-39) |
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The mistake of regarding all living cells
as membrane-enclosed dilute solutions was fully recognized by micro-anatomists
in the 1860's. Max Schultze announced his protoplasmic
doctrine in 1861.11 Thomas Huxley lectured on protoplasm as the
physical basis of life in 1868.72 Yet not until the turn of the
century did a new generation of cell physiologists begin rising to the
challenge. The research efforts of these early protoplasm-oriented cell
physiologists centered around two subjects: (i) cell swelling, and (ii) the selective accumulation of
potassium ion (K+) over sodium ion (Na+ in most
living cells—a subject already briefly introduced in [4.3]. To explain the asymmetrical distribution of K+
and Na+ in red blood cells, the supporters of the membrane theory
contended at first that the red blood cell membrane is impermeable to both
ions.76 Benjamin Moore of the University of Liverpool objected
briefly in 1906 to this impermeability interpretation.77 Two years
later, he and Herbert Roaf further elaborated on
their stand.77 First they reasoned that the content of K+
in the cell is more or less constant in the cell's life cycle. It would be very
difficult to see, they argued, how the cell could maintain a constant K+
concentration inside the cell as it grows and multiplies, if at all times its
membrane is impermeable to this ion. As an alternative, Moore and Roaf
suggested that the cell protoplasm possesses a special affinity, or
adsorbing power for K+, but it has no adsorbing power for Na+.
In support, they mentioned the preferential uptake of oxygen by living red
blood cells (erythrocytes), and, even more to the point, selective uptake of Ê+
over Na+ by (inanimate) soils. However, they did not offer a
molecular mechanism for the preferential adsorption in either the living cells
as they hypothesized, or in soils as a known fact {see [10.1(3)]}. Ñ. E. Overton, better known
for his lipoidal membrane theory (to be
discussed in [13.1(1)]), discovered in 1902 reasons to question that living
cells are truly membrane-enclosed dilute solutions. Thus when he transferred a
frog sartorius muscle from an isotonic 0.7% NaCl solution to a hypotonic NaCl
solution at half of its strength (0.35%), the muscle did not swell to twice its
initial weight as expected on the basis of the membrane theory. Instead an
increase of only a third of its initial weight was observed. Overton concluded
that at least a part of the cell water must be Quellungs-wasser
(swelling water, imbibition water).25 p
273 In 1907 and 1909 Martin Fischer, then a
professor of medicine in the Oakland School of Medicine in Oakland, California,
argued that the swelling of living cells is not an osmotic membrane phenomenon
as widely believed then [4.1], but a result of the strong affinity of
protoplasmic colloids for water—seen also in fibrin (the protein of blood
clots) and gelatin.546; 78 Along this line of reasoning, he proposed a theory of oedema and published a lengthy thesis on the subject.78
In this 1909 document he also offered briefly some new insights on the
asymmetrical distribution of ions and other solutes in cells, which the
distribution of K+ and Na+ exemplify. Fischer pointed out that the dissolved
substances may be either at higher or lower concentration in the colloidal mass
(protoplasm) than in the surrounding medium. Adsorption could then
account for its being at a higher concentration; the law of partition
(also known as the Berthelot-Nernst distribution law,420 of which Henry's law for the distribution of
gases in a liquid solvent is a special case—GL's addition) may account for its
presence at a lower concentration.78 pp 545-546
But Fischer did not further exploit these important ideas. His major scientific contribution to cell
physiology aside, Martin Fischer should also be remembered for his kindliness
and generosity, giving support out of his own pocket to even his scientific
opponents in Germany at the end of World War I.79 Quoting
Schopenhauer, colloid chemist Wolfgang Ostwald wrote
on Fisher's 60th birthday: "As torches and fireworks pale and become
invisible in the sunlight, so the mind and even genius and beauty are outshone
and overshadowed by (the) goodness of heart."79 p 441 W. W. Lepeschkin
also rejected the concept that living cells are membrane-enclosed dilute
solutions. As mentioned earlier, when he crushed young cells of the plant, Bryopsis, in sea water, many water-immiscible
little balls of protoplasm emerged62 pp 289-290—not
unlike that shown in Figure 3. On diluting the surrounding sea water with
distilled water, these little balls swelled prodigiously and vacuoles formed
within them. When these little balls were returned to sea water, they regained
their original size and the vacuoles disappeared. Lepeschkin estimated that the surface of these little balls
created by shaking could be increased a thousand fold from the surface area of
the intact cell. Because the total amount of (postulated) lipoids in the
original membrane of the intact cell is limited, it could not be expanded to
1000 times its original dimension.62 p 275
Accordingly, he concluded that these findings contradict the idea—emerging from
a synthesis of the lipoidal membrane theory of
Overton21 and the theory of membrane regeneration [4.1(4)]—that each
of these little balls is covered with a continuous lipoid membrane as
postulated by Overton for all living cells. By the same token, the surface of
these little balls must be covered with materials that are abundant in the
protoplasmic droplet as a whole, i.e., protoplasm62 p 276—in
harmony with Franz Leydig's, Max Schultze's
belief that the cell surface is made of similar materials that make up the bulk
of the cell's protoplasm. Surprising as it might seem, Wilhelm Pfeffer shared a similar view.18 p 156 In further support of Lepeschkin's
idea, one may quote the observation of Pauli and Rona
mentioned in [6.2]. They showed that a salt solution of (pure) gelatin warmed
up to 30°C forms immiscible coacervate. Since there is no other
structure-forming material than gelatin here, the membrane of the coacervate—if
one calls the surface layer of coacervate a membrane—can only be made of the
same material, which makes up the bulk of the coacervate, i.e., gelatin. As mentioned above, Lepeschkin
was among the earliest in maintaining that protoplasm is a coacervate.324
He also offered a theory of the fundamental substance of living matter, a
loose chemical complex of proteins and lipids, which he called vitaproteids or vitaids.325 This
specific hypothesis was severely criticized332; 92 p 62
and not further defended. Then there was Ross Gortner
(1885-1942), for more than a quarter of a century the Chief of the Division of
Agricultural Biochemistry at the University of Minnesota and a strong advocate
for the existence of bound water in living cells. In an address given
at the Faraday Society meeting to a group of scientists with diverse first-hand
knowledge on various aspects of water—the printed version of which bears the
title: "The State of Water in Colloidal and Living Systems" and was
published in the Transactions of the Faraday Society80—Gortner
presented his view that at least a part of the water in living cells is not
normal liquid water and is referred to as bound. As one of the criteria that
sets bound water apart from normal water, Gortner
suggested that bound water does not dissolve substances soluble in normal
liquid water—hence an alternative name for bound water in his and his
coworkers' vocabulary is "non-solvent water." In anticipation of what follows in Chapter 11, I mention
that Gortner referred to experimental evidence in
the literature for the existence of multilayers of adsorbed water in inanimate
systems. However, Gortner then backed away in these
words: "Unfortunately the properties of water in oriented adsorption
films have not been sufficiently characterized to enable us to state whether or
not this may be the type of water which the biologist is coming to call bound
water."80 pp 684-685 Gortner's paper presented at the Faraday Society meeting was
well received. A majority of the participants expressed interest, some very
enthusiastically. However, there was one notable exception, i.e., the same
Professor A. V. Hill, whose evidence for free cell water has already been
briefly presented in [5.1]. Not mentioned earlier is the fact that A. V. Hill
almost single-handedly extinguished the enthusiasm generated for the
"bound water" concept, and put to rout the entire colloidal approach
to cell physiology. He achieved this feat by presenting just a single set of
experimental data demonstrating that water in frog muscle accommodates the same
concentration of urea as in the surrounding fluid.81 Accordingly, there is no "nonsolvent"
water. (In years following, others showed that ethylene glycol also
distributes equally across the cell membrane of rectus
muscle and of red blood cells.82) Since cell water is thus proven to
be normal liquid water, the components in the cells high enough in
concentration to match osmotically the free Na+ and Cl- in a Ringer's solution, can only be free K+
and free cell anion(s). Ergo, no bound water, nor bound K+ in living cells81—in affirmation of the
clinching twin tenets of the membrane theory. Hungarian biophysicist, Professor E. Ernst, a witness
to these events, recalled how the opinion makers of the day, including Rudolf Höber (despite his own electric conductance study
pointing to the contrary [5.2]), W. O. Fenn, and F. Buchthal all renounced the concept of bound water and bound
K+, and subscribed in toto to the idea that
living cells are membrane-enclosed dilute solutions following van't Hoffs osmotic law.83 p
112 Each acknowledged that his new position was based on Hill's
crucial experimental finding of equal urea distribution and his compelling
logic. In 1940 the major English-language Journal of Colloidal Chemistry was combined with the Journal of Physical Chemistry. For a few years, the combined journal bore the new title Journal of Physical and Colloidal Chemistry. Then the words "and Colloidal" were quietly dropped. The demise of the main English-language colloid magazine did not signal the end of colloid chemistry per se. Other journals like Zeitschrifts für Kolloid Chemie, Kolloid Beihefts and even Protoplasma continued publication. It was only the fledgling colloid- or protoplasm-oriented cell physiology that suffered an (undeserved) near-fatal blow. |
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