3. CELLULAR ASPECTS OF ACTIVE TRANSPORT
149
define the exact nature of the forces involved in this concentration
process. The main obstacle arises from the fact that we have little, if
any, control on the intracellular phase. The various parameters defined
in Section II are not experimentally accessible, and some hypothesis
must be made as to the intracellular physicochemical state of the compound under study. It has been shown that glycine could be 20 times
as concentrated in the interior of the isolated cells as in the surrounding
medium (12). If glycine is free in the cell, its concentration must be
the result of an active transport. At the physiological pH, glycine is
indeed mostly a molecule without net electric charge, a small part of it
being anionic; the only important parameter is then the concentration,
or more precisely, the activity. If, however, glycine is bound, it could
enter the cell passively, bind some cytoplasmic constituent, and thus
simulate an active transport. A distinction between these two modes of
entry is not possible from steady state distribution or from the simple
observation that the transfer is dependent upon metabolic process. The
binding could indeed be endergonic.
It has been assumed on the basis of various observations that most
of the intracellular amino acids are free. Fragments of cancer cells
obtained by sudden pressure release (50), or by grinding after freezedrying (51), are unable to accumulate amino acids. Accumulation of
amino acids at high concentration was found to disturb the osmotic
equilibrium between intra- and extracellular fluid (11). As will be discussed in more detail in a following section, amino acids are an important constituent of the osmoregulatory mechanisms in invertebrates,
suggesting that they are osmotically active and thus really free. The
loss of radioactivity from cells previously incubated with labeled glycine
is not enhanced on addition of large amounts of unlabeled glycine
to the medium (51). These observations, if they do not prove, at least
strongly suggest, that amino acids are free in the cell interior. More
direct and conclusive evidence has been published by Heinz (52) and
Oxender and Christensen (5). The experiments of Oxender and Christensen have already been discussed above. Heinz has produced more
direct information as to the nature of the forces involved in the transfer
process of glycine in the Ehrlich mouse ascites cells by studying the
kinetics of the fluxes with and without inhibition. He was able to show
that cells in the steady state with respect to the distribution of glycine
exchange more than 90% of their glycine in about 5 minutes. The kinetics
of this exchange seems to indicate that intracellular glycine behaves as
though most of it is freely dissolved in a single compartment. It can
therefore be concluded that the accumulation within the cell is the effect
of a metabolically linked active transport mechanism.
149
define the exact nature of the forces involved in this concentration
process. The main obstacle arises from the fact that we have little, if
any, control on the intracellular phase. The various parameters defined
in Section II are not experimentally accessible, and some hypothesis
must be made as to the intracellular physicochemical state of the compound under study. It has been shown that glycine could be 20 times
as concentrated in the interior of the isolated cells as in the surrounding
medium (12). If glycine is free in the cell, its concentration must be
the result of an active transport. At the physiological pH, glycine is
indeed mostly a molecule without net electric charge, a small part of it
being anionic; the only important parameter is then the concentration,
or more precisely, the activity. If, however, glycine is bound, it could
enter the cell passively, bind some cytoplasmic constituent, and thus
simulate an active transport. A distinction between these two modes of
entry is not possible from steady state distribution or from the simple
observation that the transfer is dependent upon metabolic process. The
binding could indeed be endergonic.
It has been assumed on the basis of various observations that most
of the intracellular amino acids are free. Fragments of cancer cells
obtained by sudden pressure release (50), or by grinding after freezedrying (51), are unable to accumulate amino acids. Accumulation of
amino acids at high concentration was found to disturb the osmotic
equilibrium between intra- and extracellular fluid (11). As will be discussed in more detail in a following section, amino acids are an important constituent of the osmoregulatory mechanisms in invertebrates,
suggesting that they are osmotically active and thus really free. The
loss of radioactivity from cells previously incubated with labeled glycine
is not enhanced on addition of large amounts of unlabeled glycine
to the medium (51). These observations, if they do not prove, at least
strongly suggest, that amino acids are free in the cell interior. More
direct and conclusive evidence has been published by Heinz (52) and
Oxender and Christensen (5). The experiments of Oxender and Christensen have already been discussed above. Heinz has produced more
direct information as to the nature of the forces involved in the transfer
process of glycine in the Ehrlich mouse ascites cells by studying the
kinetics of the fluxes with and without inhibition. He was able to show
that cells in the steady state with respect to the distribution of glycine
exchange more than 90% of their glycine in about 5 minutes. The kinetics
of this exchange seems to indicate that intracellular glycine behaves as
though most of it is freely dissolved in a single compartment. It can
therefore be concluded that the accumulation within the cell is the effect
of a metabolically linked active transport mechanism.