ROUND TABLE DISCUSSION 769 
mind, however, that the formation of an activated complex such as that between acetaldehyde 
and thiaminpyrophosphate, involves a highly specific and energy-rich covalent bond. There may 
be plenty of sites available inside the cell, which could be imagined to function as more less un- 
specific adsorption sites. I find it difficult, however, to visualize enough specific sites able to form 
activated complexes with the above-mentioned characteristics, in order to accomodate the tre- 
mendous amounts of amino acids accumulated. 
Furthermore, a similar dependence of a unidirectional flux on its transconcentration has later 
been observed with other systems where such an activated binding was strictly excluded. So we 
found with DurRBIN® that the chloride flux across the gastric mucosa towards the lumen is also 
increased by the chloride tvansconcentration. A counter-flow effect was also reported by Park 
et al, and by ROSENBERG AND WILBRANDT!" with the flux of certain sugars out of the red cell. 
Here the tvansconcentration, contrary to our system, referred to the extracellular concentration 
of the substrate which was undoubtedly in a free dissolved state. Of special importance are prob- 
ably the more recent experiments of Dr. CHRISTENSEN ef al. which reproduced the preloading or 
counter-flow effect between two extracellular compartments, which were separated by a layer of 
ascites cells arranged to a solid membrane. So I think that by analogy, the preloading effect as 
originally observed is strongly suggestive that the crucial amount of accumulated amino acids in 
our cells is in a free dissolved state. By this, of course, I do not mean a state equal to that of an 
ideal gas but a state comparable to that of the extracellular amino acid. I have to concede that a 
certain extent of intracellular adsorption more or less loose and unspecific cannot be rigidly ex- 
cluded. Such adsorption would, in our view, not be metabolically linked and hence rather inci- 
dental with respect to the active accumulation mechanism. 
In this connection I would like to draw your attention to another phenomenon which we 
reported several years ago (HE1Nz!*). If a cell has accumulated a certain amino acid, for example 
glycine, then—according to the CARNEGIE foundation—I would assume that there would be a 
concentration or activity of the free amino acid similar to that in the medium, and in addition, 
there would be a certain amount of glycine which is what you call bound. Is that right? 
CowleE: Yes. 
Heinz: This bound amount is with normal cells. If you now do the same experiment, but add an 
efficient metabolic inhibitor as DNP, then you get, of course, the same amount of free amino acid 
as in the uninhibited control. Owing to this inhibitor much less binding could be expected. 
If we now measure the initial rate of efflux in this case, it should be proportional to the concen- 
tration (or activity) of the free glycine, and not of the total glycine. Does this agree with your 
views? 
Britten: I think I indicated that the efflux in such a case would be proportional to the number 
of loaded carriers and their dissociation constant. This is a free parameter, so that I can adjust it 
to meet the results of your experiment. That is, there are circumstances in which you may get very 
slow losses, and others in which you can get relatively rapid ones, depending on the nature of the 
carrier. This carrier has taken a great burden of properties. 
He1nz: We actually found that the efflux, with and without inhibition, was almost proportional 
to the total cellular glycine. In other words, metabolic inhibition or anoxia reduced the steady 
state efflux by a factor of up to six or seven. Accoiding to Dr. BriTTEN’s statement, that the efflux 
is “proportional to the number of loaded carriers”, our finding would indicate that in metabolic 
inhibition this number of loaded carriers is drastically reduced, e.g. 6—7 times. In Dr. BRITTEN’s 
carrier model, however, it is not the loading of the mobile carrier which is coupled to an energy 
donor, but the transfer of the amino acid from this carrier complex to the non-mobile site—7.e. a 
reaction which clearly tends to decrease the number of loaded carriers, whether they are saturated 
or not. This model would, therefore, predict that metabolic inhibition, if anything, increases rather 
than decreases the efflux. As already mentioned we found precisely the opposite. Provided that our 
results give the true steady state efflux values, and provided that the metabolic inhibition is 
specific enough, these results support the view that the accumulation of amino acids is brought 
about by an active transport process and not by an endergonic binding to non-mobile sites. I do 
not see how one could adjust the parameters of Dr. BRITTEN’s model to meet these results except by 
the assumption that either these parameters or the passive permeability of the cell membrane are 
profoundly altered by metabolic inhibition or anoxia, an assumption, which to my knowledge is 
not made in your model. I wonder whether Dr. REINER would like to comment on this point. 
REINER: I think that theoretical models may appear to be fundamentally different—judging 
by the elements that are involved in them. That is, a pump or a membrane model can be made to 
give varied predictions. By changing some relationships in this—and I think this could quite 
possibly be done—you could perhaps make its predictions come out to be very much like the predic- 
tions of the carrier model. In fact, this philosophical problem of distinguishing the object that you 
are talking about from the mathematical interpretation of the quantitative aspects of an experi- 
ment is extremely difficult. To be resolved it needs a crucial experiment, and crucial experiments 
very often get denuded when you analyze them. 

References p. 777 
