Temperature Dependence of DF2’s Effectiveness is Due to Temperature Sensitivity of Phosphatases
The neuropeptide DRNFLRFamide (DF2) increases synaptic transmission at the crayfish neuromuscular junction, but its function depends on temperature. The difference in DF2’s effectiveness in synaptic modulation may be due to different degree of dephosphorylation by protein phosphatases. Motivated by previous studies on temperature dependence of synaptic enhancement induced by DF2, we hypothesized that (1) DF2’s function depends on phosphorylation and inhibition of phosphatases would increase DF2’s effectiveness. (2) At lower temperature phosphatases will be less active. Therefore, increase in DF2’s function at low temperatures is due to more phosphorylated proteins induced by inactiveness of phosphatases. We experimented with DF2 and phosphatase inhibitors at both low and room temperatures. We determined the effectiveness of DF2 by measuring Excitatory Postsynaptic Potential (EPSP) of extensor muscle cells in different solutions with five minutes of adjustment period using intracellular recording and repeated stimulation of every five seconds. Our data shows that DF2’s effectiveness increases at low temperature, and the effect of phosphatase inhibitors at low temperatures was not as prominent as that at room temperature. The results indicate that at low temperature, phosphatase has already been inhibited, and further inhibition with phosphatase inhibitors has no notable effect. This result supports our hypotheses, suggesting that the effectiveness of DF2 increases in low temperature due to minimized function of phosphatase.