All of these theories fall into two groups: those which consider the action of the anesthetic to be upon the cell-membrane and those which consider the action to be directly upon the cell-contents. Experiments on the anesthesia of light-production in Nodituta have been carried out in the attempt to find out which of these two alternatives is correct.
Although the anesthesia of some processes has been shown to be independent of the consumption of oxygen—for instance, the cleavage of sea-urchin eggs (Loeb and Wasteneys, 1913, and Warburg, 1910)— it would seem probable that the anesthesia of light-production is depen dent on oxygen. That oxygen is necessary for light-production in Noctiluca has been shown by keeping the cells in an oxygen-free atmos phere, in which case the luminescence stops, but reappears on admitting oxygen. The membrane is freely permeable to oxygen at any time, since those cells deprived of oxygen immediately glow on admission of oxygen. It may be noted in passing that this oxidation is different from that of other cells, as shown by the fact that RCN in relatively high concen trations has no effect, whereas it so quickly affects other oxidations.
The anesthesia must in some way affect the oxidation which is con cerned in light-production. Does it do so by preventing oxygen from entering the cell, or does it prevent the utilization of the oxygen already present in the cell? In other words, does the anesthetic act upon the membrane or directly upon the cell interior? In my experiments various lipoid-soluble anesthetics were tried and the effect of the anesthetic has been in all cases not to prevent light production altogether, but to prevent a normal response—i. e., a flash ing on stimulation. In all the effective concentrations, the animals under the anesthetic produce a steady glow, so faint in some cases that it is not noticeable unless the animals are present in large number. When returned to sea-water, if not left too long in the solution, the steady glow ceases and the normal response returns; this is therefore a reversible phenomenon and a true case of anesthesia. The best concen trations for anesthetizing, as seen by referring to table 6, were: 1/3 sat urated chloroform, where the steady glow lasted 2 hours; m/8 ether and m/8 butyl alcohol, steady glow lasting 1 hours; 1/16 saturated thymol, lasting 1 hour; methyl alcohol, lasting 30 minutes, and 1/4 to 1/8 chloretone, lasting 15 minutes. If returned to sea-water after the period of steady glow, the animals gave no response, the prolonged anesthesia causing death. The tentacle motion was also stopped by the anesthetic in the same concentrations as prevented the normal light-response. The effect on the tentacle was, however, much slower than the effect on light-production, but took place during the early part of the period of constant glow.
The question as to whether the effect of the anesthetic is upon the interior of the cell or upon the cell membrane was tested by destroying membranes, thus allowing oxygen to enter. A number of noctilucas were subjected to m/8 butyl alcohol for an hour in a tube containing sand. Some were then returned to sea-water and recovered, showing that the animals were truly narcotized. The remaining animals were shaken up and broken to pieces by the sand. At the same time, a control lot kept for the same period in sea-water in a tube containing sand were shaken and broken up. A comparison of the two tubes showed that the anesthetized animals were not nearly so bright as the control lot. A microscopic examination of the material showed that the cells were completely broken to pieces into irregular fragments by the sand. A similar experiment was tried with animals subjected to m/8 ether for 10 minutes with the same result. In these experiments the cell-mem branes were destroyed, allowing oxygen to enter the cell directly, and the effect was different for the anesthetized cells and the control. The anesthetic must therefore act, not on the cell-membrane by preventing oxygen from entering the cell, but directly upon the cell interior, prob ably attacking the mechanism of the utilization of oxygen in the cell.
A criticism of the foregoing experiments might be offered—that, owing to the length of time that the cells were subjected to the anes thetic, substances present in the cells might be used up and the cells be fatigued. The following experiment showed that such was not the case: Three lots of animals were put into tubes with sand, two im mersed in m/8 ether and the third in sea-water. One of the ether tubes was very slightly agitated almost immediately and did not increase in brightness as normal animals do on slight agitation, showing that the animals were narcotized. The other ether tube was shaken vigorously at once so as to break up the animals, and it became slightly brighter. The control tube with sea-water, when also shaken vigor ously at the same time, became markedly brighter, showing a striking contrast to the other two tubes. In this experiment there was not sufficient time for any substances in the cells to be used up, and yet the narcotized cells gave a much weaker light than the control lot on admission of oxygen by destroying the membranes. The evidence from these experiments on Noctiluca would therefore argue against the mem brane theory of anesthesia, since the anesthesia of light-production takes place independently of the cell-membrane; the effect of the anesthetic seems to be upon the mechanism of the oxidation process inside the cell.