It’s an interesting question, Jeremy, why those tests were done as they were, and not on skeletal muscle.
In medicinal chemistry, one of the main things that is done is that when one has a “lead compound,” some compound that has a desired activity, one makes some derivatives and analogues of it – changed versions of it – that are strategically planned so as to be likely to perhaps be better, and also to have synthetic organic chemists make perhaps 1000 or more other analogues and derivatives, more or less randomly according to what they happen to be able to make. So you now you have 1000 or more candidate molecules of which perhaps some are effective drugs having one or more useful advantages over the original lead compound.
What’s required next is an “assay” to pick out which of these compounds show promise. Ideally, this would be some simple test-tube experiment. For example, if you are designing a drug to be an enzyme inhibitor, it’s easy to determine in a test tube how well a compound inhibits the enzyme, so all 1000+ compounds can be screened quickly and inexpensively. Let’s say 50 look promising. The next step would be to run perhaps a mouse or rat assay and see how well the inhibitor works in the living animal. This is a more expensive step of course. Based on these results and further toxicological studies, and probably on other concerns such as formulations and stability, ultimately perhaps one of these 1000+ compounds you will deem suitable for human trials.
In the case of anabolic steroids, there was no test tube test. So the question was, how to test 1000 or more compounds in rats cheaply and quickly.
But test them for what?
What scientists were trying to do was what you always try to do if your lead compound has more than one activity. You try to develop analogues and derivatives that have only one of those activities, or at the very least, are more specific than the lead compound was. Scientists were hoping that the protein anabolic and the virilizing effects of testosterone were separate properties and potentially drugs could be developed that would have only, or at least mostly, just one or the other.
So, what was needed was a means to assay
potency both of something that was hoped to
be an indicator of potency in protein anabolism, and another that was hoped to be an indicator of potency in virilization.
A scientist named Hirshberger came up with the idea of injecting rats with anabolic steroids, then measuring growth of two of the very few organs which respond rapidly to androgens, namely the levator ani (which corresponds to the pubococcygeus or “PCG” muscle in humans) and the prostate. The first was hoped to be a good predictor of potency in protein anabolism and the second a good predictor of potency as a virilizing agent.
This assay was easily performed and different drugs did show a “better” ratio of activity than testosterone did, so the test was thought to be a success and well over 1000 compounds were screened that way.
Only problem is we now know that what was measured was simply potency in the levator ani of the rat and potency in the prostate of the rat. Not that much else was well predicted from these tests.
A Russian scientist developed a better assay involving cutting the gastrocenemius muscle of the rat, thus forcing the soleus (or perhaps it was vice versa) to do all the work as the rat was forced to run on a treadmill. This was a good assay but unfortunately was never widely adopted.