SUMMARY1. In denervated guinea-pig diaphragm the depolarization produced by decamethonium (100 pM) was followed by an initial phase of recovery, and then by a slow restoration ofmembrane potential in the presence ofthe drug, with hyperpolarization. Membrane potentials were measured by repeated insertions. The slow phase of spontaneous recovery was not found in the absence of potassium or in the presence of ouabain (100/M).2. With 1 /SM-decamethonium the net loss of potassium from denervated muscle was 17 % by wet weight in 20 min as compared with controls, which represents a loss of over 30 mm in internal concentration. Similar results were obtained with 100 FMdecamethonium. Spontaneous recovery of potassium occurred in the succeeding 2 h in the presence of 1 /SM and 100 /SM-decamethonium. With 5 nM-decamethonium muscles exposed for 20 min had a potassium content which was not reduced as compared with controls.3. In rat diaphragm decamethonium (100/M) also produced depolarization and slow spontaneous recovery which was not seen in the absence of potassium or the presence of ouabain. With 3 mM-decamethonium spontaneous recovery of potential was complete in 5 min.4. Change from 5 mM-potassium to potassium-free solution produced consistent hyperpolarization in denervated guinea-pig diaphragm. In rat diaphragm at 38 0C the results were variable, with some fibres showing hyperpolarization while others showed depolarization.

1 The water content and mannitol space of rat diaphragms which were perfused through the inferior vena cava was increased compared with immersed diaphragms. The potassium content of both preparations, when expressed in terms of dry weight, was maintained at similar values to that found in vivo. 2 Despite the application of a constant concentration of decamethonium, a steady level of neuromuscular block was not obtained in either the perfused or immersed rat diaphragm. The immersed preparation differed from the perfused preparation in that recovery from paralysis occurred despite the continued presence of the drug. 3 The rate of uptake of labelled decamethonium (100 Mm) at the end-plate region was similar in the perfused and immersed diaphragm. The slopes of the regressions were 0.059 and 0.054 ,ul mg1min-' (based on dry weight) respectively which were not significantly different. This implies that the rate of uptake of the drug at the end-plate is slow and limited by the rate of entry into the fibre rather than by diffusion to the site of entry. IntroductionMethods Creese & Maclagan (1970) have obtained autoradiographic evidence that decamethonium enters muscle fibres in the rat. It has been suaggested that the permeability of this organic ion at the end-plate is low and similar to that of sodium (Creese & England, 1970). If this is the case, then the uptake of decamethonium would be expected to be limited by entry at the cell membrane rather than by diffusion. If however, the permeability of decamethonium approached that of potassium then the uptake would be faster in a perfused muscle than in an immersed muscle because of delays due to diffusion in the latter (Klaus, Luilman & Muscholl, 1960;Keynes, 1954). In the present study, rat diaphragms were perfused in vitro via the inferior vena cava with labelled drug to compare the rate of uptake with that in immersed muscles. In addition, the neuromuscular block produced by decamethonium was recorded in an attempt to confirm previous claims that a steady paralysis can be obtained with depolarizing drugs (Maclagan, 1962; Gibberd, 1966) when applied via the capillaries. The water, potassium and sodium content and also the mannitol space were determined in both preparations. Immersed diaphragmThe left anterior portion of the diaphragm together with its rib was quickly removed, with or without the left phrenic nerve and transferred to a bath (100 ml) for recording of contractions, or to a tube (10 ml) for measurement of uptake of labelled decamethonium. Twitch tension was recorded by means of a strain gauge connected to the tendon by a thin platinum wire (0.4 g). The uptake of the labelled drug was measured in the manner described by Creese & England (1970).After immersion in labelled drug the muscle was 368 P.P.A. HUMPHREY washed in saline for 10 min, attached to a metal strip and frozen on solid carbon dioxide. Perfused diaphlragmThe preparation is similar to that described by Gibberd (1966). The inferior vena cava was cannulated through the right auricle and the e...