Fig. 8. Tyrosine phosphatase inhibition causes the accumulation of recycled receptors in the peri-synaptic membrane. (A) Live adult mouse NMJ previously saturated with BTX-biotin/strept-488 (green) at time 0 and viewed at high detector gain 9 hours later. The muscle was continuously bathed in the tyrosine phosphatase inhibitor PAO for 9 hours and then imaged. (B) The junction was then incubated with strept-594 (red) to determine if PAO affects AChR recycling. Perisynaptic areas were labeled (arrows), indicating that some of the recycled receptors had become mistargeted. (C) Overlay of pre-existing and recycled receptors (overlay images were adjusted for brightness and contrast using Photoshop). (D-F) Similar results were obtained using an alternative tyrosine phosphatase inhibitor, pervanadate. Note that whereas recycled receptors (red) appear in the peri-synaptic area, pre-existing receptors (green) remain restricted to the synapse. (G-I) When the serine/threonine phosphatase inhibitor okadaic acid was used instead, recycled receptors (red) were properly targeted to the synapse. Scale bars: 20 µm.