The cellular prion protein (PrP) plays important roles in many neuronal processes including memory, circadian rhythm, neuroprotection and neurotoxicity1. Although physiological roles of PrP remain elusive, its neurotoxic signalling has been investigated more thoroughly. As a consequence of PrP misfolding, several mammalian species develop neurodegenerative conditions known as scrapie, bovine spongiform encephalopathy or Creutzfeldt-Jacob and Gerstmann-Sträussler-Scheinker syndrom (GSS) in human. The current study investigated synaptic effects of expressing mouse PrP and a mutated form of PrP (PrPP101L, resembling a GSS-like disease in mice) in Drosophila to elucidate synaptic functions of PrP. The UAS/Gal4 bipartite expression system was used to drive pan-neuronal expression with the elav-GAL4 driver2. Two-electrode voltage clamp experiments were conducted to record miniature and evoked excitatory junctional currents (m/eEJCs). Intracellular recordings from larval muscle 6 were performed (Milticlamp 900A, pClamp 10, Axon Instruments) in haemolymph-like solution 3 at 25oC. Data are expressed as mean±SEM (n). Statistical analyses were carried out using ordinary ANOVA and Kolmogorov-Smirnov (K-S) tests, * indicates p<0.05. PrP expression induced an increase in mean mEJC from 0.77±0.03nA (31) in wild type (WT) to 1.17±0.12nA* (4). This augmentation was also observed following expression of PrPP101L (0.92±0.04nA* (17)) although to a lesser extent. The increase in mEJCs is due to a shift in the distribution of single mEJC amplitudes. Relative cumulative frequency histograms for mEJC amplitudes showed an increased probability of larger amplitudes and hence a right shift in mEJC distributions in PrP expressing larvae (K-S test: PrP: D=0.28, p<0.0001, PrPP101L: D=0.24, p<0.0001) relative to WT. mEJC decay (τ) values did not differ between genotypes (τWT=10.0±0.4ms; τPrP=12.1±1.3ms; τPrPP101L=10.8±0.4ms) nor did τ distributions, indicating that postsynaptic receptor composition was not affected. The frequency of mEJCs was not different between the genotypes (fWT=1.6±0.2s-1; fPrP=1.6±0.4s-1; fPrPP101L=1.3±0.2s-1). In agreement with electrophysiological data, electron microscopy revealed that PrP and PrPP101L (but to a smaller degree compared to PrP) expression caused an increase in vesicle diameters (1s boutons: WT: 43.4±0.7nm (6); PrP: 55.4±2.3nm* (3); PrPP101L: 47.5±0.4nm* (4)) suggesting that larger vesicles are responsible for the augmented mEJC amplitudes. Further, PrP expression caused an increase in foraging behaviour as detected in greater crawling distances per 30min (WT: 1.2±0.1m (23); PrP: 1.9±0.1m* (23); PrPP101: 1.8±0.1m* (13)). This data indicate that PrP mediates some of its physiological effects via formation of synaptic vesicles and that mutated PrP (PrPP101L) as present in prion disease loses this ability thereby contributing to disease progression on the synaptic level.