Acetylcholinesterase and the acetylcholine receptor are two postsynaptic proteins which play an important role in neuromuscular transmission. They are both affected by muscle denervation. Extra-junctional membrane which is normally insensitive to acetylcholine increases its sensitivity after denervation, and in mammalian muscle acetylcholinesterase activity decreases. The proposed experiments will attempt to study both the biochemical properties of these two proteins, and the mechanisms by which their levels are regulated in normal and denervated muscle. Our studies on acetylcholinesterase in intact tissue and crude homogenates have suggested the hypothesis that the 16 S form of the enzyme is specifically associated with the endplate, and that this form is removed by collagenase or trypsin and converted to a 10 S enzyme. To investigate this problem, we propose to purify the various forms and examine the effect of proteolytic enzymes on each species. We will further test the proposition that the 16 S form is the endplate enzyme by trying to determine whether each of the various forms is on the interior or exterior of the muscle fiber. In our studies on the acetylcholine receptor protein, we will first try to define the pharmacological properties of the isolated receptor. Since our experiments on loss of toxin from muscle in vivo suggested that there might be differences between normal receptor and those which appear after denervation we will attempt to purify each of these and compare their properties. The regulation and turnover of these two proteins will be investigated by using organ culture systems and whole animal preparations. The effect of muscle activity on acetyl-cholinesterase activity will be determined both by direct electrical stimulation of denervated muscles and by interrupting neuromuscular transmission in innervated muscles. Methods will be developed for directly measuring synthesis and removal of the acetylcholine receptor and the relative rates of these processes determined both in physiological situations where extra-junctional receptor concentration is increasing and in situations where it is decreasing.