Notch is required for long-term memory in Drosophila

Abstract

A role for Notch in the elaboration of existing neural processes is emerging that is distinct from the increasingly well understood function of this gene in binary cell-fate decisions. Several research groups, by using a variety of organisms, have shown that Notch is important in the development of neural ultrastructure. Simultaneously, Presenilin (Psn) was identified both as a key mediator of Notch signaling and as a site of genetic lesions that cause early-onset Alzheimer's disease. Here we demonstrate that Notch loss of function produces memory deficits in Drosophila melanogaster. The effects are specific to long-term memory, which is thought to depend on ultrastructural remodeling. We propose that Notch plays an important role in the neural plasticity underlying consolidated memory.

Fig. 1.

Conditioned courtship indicates that Notch dysfunction impairs long-term memory. (a) Short-term memory was examined 30 min after training. Wild-type (CS) and N^ts1 flies display intact short-term memory after 2 days in nonpermissive conditions.^*, Statistical differences between naïve and trained cohorts indicating memory of training (P = 0.0087 for CS and P = 0.0088 for N^ts1). (b) Long-term memory was examined 2 days after training. CS males show intact long-term memory (P = 0.0007 for CS) after 2 days in nonpermissive conditions, whereas N^ts1 flies do not demonstrate long-term memory. (c) Long-term memory was examined in flies maintained at permissive temperatures to test for developmental enfeeblement. Under permissive conditions, both CS and N^ts1 flies display statistically significant differences in naïve versus trained data sets, demonstrating the presence of long-term memory (P = 0.0188 for CS and P = 0.0003 for N^ts1). (d) Two-day long-term memory was examined in N^ts1 flies bearing heat shock-driven wild-type Notch transgene (N^ts1;hs-N⁺/+) or a phenotypically nonfunctional control Notch missing ligand-interaction domains (N^ts1;hs-ΔEgf-Ni/+). Only flies carrying functional Notch displayed statistically significant rescue of long-term memory (P = 0.0430 for N^ts1;hs-N⁺/+). These results indicate that requirements for long-term memory are specific for functional Notch. At least 15 flies were examined in every cohort shown.

Fig. 2.

N^ts1-mediated Notch dysfunction impairs long-term memory, as demonstrated by Pavlovian olfactory conditioning. (a) Short-term memory was examined 30 min after training in flies kept in nonpermissive conditions for 4 days. Flies displayed strong avoidance of the odor paired with footshock, demonstrating intact short-term memory (P < 0.0001 for both). There were also no statistically significant differences between behavior of CS and N^ts1 flies. (b) Flies were kept in nonpermissive conditions for 2 days, were trained, and were then tested for long-term memory 1 day after training. Under nonpermissive conditions, CS flies displayed statistically significant odor avoidance (P = 0.0182 for CS), whereas N^ts1 fly performance was impaired and did not differ significantly from 0 (indicated by an asterisk). These results indicate that N^ts1 flies have an impaired long-term memory. (c) Long-term memory was examined in flies maintained at permissive conditions. Both CS and N^ts1 flies displayed significant odor avoidance, indicating that long-term memory is intact for both (P = 0.0003 for CS and P = 0.0005 for N^ts1). Data for female flies are shown above. We found that females performed slightly better than males under these nonoptimal conditions for memory testing, although males showed the same trends. Data from six PIs for short-term (representing ≈1,200 flies) and 12 PIs for long-term memory are shown.

Fig. 3.

RNAi-mediated Notch silencing impairs long-term memory of Pavlovian olfactory conditioning. (a) Flies were examined for short-term memory 30 min after training at 25°C. We tested flies containing the responder only [UAS-Ni/+ (Ni)] as a control for insertion-site effects, flies expressing Notch RNAi in adult CNS including MBs [c772/+; UAS-Ni/+ (c772-Ni)], and flies bearing a control RNAi-producing transgene driven by the c772-Gal4 line [c772/+; UAS-LacZi/+ (c772-LacZi)]. All groups showed statistically significant short-term memory with no differences among groups (ANOVA, F₍₃₃₎ = 0.92, P = 0.4187). (b) Long-term memory was tested in Ni, c772-Ni, and c772-LacZi flies 1 day after training. Statistical analysis demonstrated long-term memory in all cohorts, but significant impairment of long-term memory in c772-Ni flies was identified by a Student–Newman–Keuls test (ANOVA, F₍₃₃₎ = 4.20, P = 0.0207, shown by an asterisk). Data from 6 PIs in short-term memory and at least 16 PIs in long-term memory are shown.