Effects of oxidative stress and aging on nerve, muscle, and synapse in a male-specific abdominal neuromuscular junction in Drosophila

Defects in Drosophila Cu ²⁺ /Zn ²⁺ superoxide dismutase (encoded by the gene Sod1 ) lead to elevated oxidative stress and a drastically shortened lifespan. To contrast the effects of aging and oxidative stress on nerve conduction, synaptic transmission, and muscle excitability, we developed an easily accessible adult abdominal neuromuscular preparation, utilizing the male-specific Muscle of Lawrence (MOL) in Drosophila . The large size of MOL facilitated analyses of presynaptic nerve signals and postsynaptic responses that could result in sizable excitatory junctional potentials (EJPs) evoking full-blown muscle action potentials (APs) which were terminated rapidly by a characteristic afterhyperpolarization (AHP). Aged wild-type (WT) individuals (80 days or older) exhibited diminished neuromuscular transmission, mainly reflecting declines in motor axon conduction, with synaptic transmission remaining largely intact (since robust EJPs could still be evoked when nerve terminals were directly stimulated electrotonically). Additionally, muscle APs showed enhanced depolarizing peaks and weakened AHPs during current injection, suggesting weakening in repolarizing K ⁺ currents. Chronologically younger Sod1 mutants (up to 30 days) displayed similar trends of neuromuscular changes, confirming a major role of oxidative stress in aging. However, certain distinctions exist in muscle membrane properties and transmitter release machinery. A clear increase in muscle membrane resistance was seen in Sod1 but not in aged WT. Additionally, unlike normal spontaneous release of synaptic vesicles leading to miniature EJPs (mEJPs), extremely enlarged spontaneous transmitter discharges occurred in aged WT but was never seen in Sod1 , indicating a distinct, aging-specific alteration in transmitter release regulation. Notably, our work revealed considerable variation among individuals, ranging from transmission failure to largely intact neuromuscular functions, demonstrating the stochastic nature of functional declines due to aging and oxidative stress. Moreover, this study uncovered a well-defined common vulnerability, i.e. weakening of the Ca ²⁺ -activated BK current that caused drastic reduction in AHP in both aged WT and Sod1 mutants, as confirmed by their diminishing sensitivity to the BK channel blocker paxilline, which caused striking alterations in the AHP in WT control.