The A-type potassium current (I_KA), mediated by Kv4 subunit containing channels, defines how cerebellar granule neurones (CGN) respond to excitation (An et al. 2000). Although tightly regulated, the functional expression of this physiologically important current is enhanced in CGN incubated with exogenous amyloid β protein 1-40 (Aβ; Ramsden et al. 2001). Aβ is constitutively derived from amyloid precursor protein by β- and λ-secretase activity and augments I_KA via an increase in protein synthesis (Plant et al. 2002). Here we use inhibitors of β- and λ-secretase to prevent endogenous Aβ production in CGN and investigate the importance of this peptide to I_KA.

Rat cerebellar granule neurones were isolated and cultured from tissue obtained by Schedule 1 methods as previously described (Ramsden et al. 2001). For whole-cell patch-clamp recordings, extracellular and pipette solutions used were those detailed in Ramsden et al. (2001). Cells were held at -70 mV and K⁺ currents (I_K) evoked by step depolarisations to +50 mV following a prepulse to -140 mV. Western blot analysis of 10 % SDS-PAGE gels was carried out by probing membranes using polyclonal antibodies raised against Kv4.2 and 4.3 α-subunits. All values are given as means ± S.E.M.

In control, untreated granule neurones the mean peak I_K measured at +50 mV was 1.2 ± 0.03 nA pF^-1 (n = 17). This was significantly reduced to 0.58 ± 0.08 nA pF^-1 (n = 17, P < 0.01, Student’s unpaired t test) following 24 h treatment with 2-Naphthoyl-VF-CHO (λ-IV), an inhibitor of λ-secretase activity. I_K was further diminished (0.36 ± 0.05 nA pF^-1, n = 15) following 48 h treatment with λ-IV. As λ-secretase activity may contribute to other signalling pathways, the experiment was repeated with H-KTEEISEVN-stat-VAEF-OH (βSI), an inhibitor of β-secretase activity. 24 h treatment with βSI resulted in a decrease in I_K which was similar to that recorded following λ-IV treatment. This suggests that these structurally distinct and secretase selective inhibitors mediate a reduction in I_K via the same pathway. In order to confirm the importance of Aβ to the functional expression of I_K we attempted to rescue the reduction in current caused to λ-IV by co-incubating with 1 nM Aβ. This treatment was effective in negating the deficit in I_K attributed to diminished Aβ production (I_K = 1.4 ± 0.05 nA pF^-1, n = 12).

These data demonstrate for the first time the importance of endogenous Aβ in determining I_K in central neurones, further supporting the contention that Aβ has a physiological role in the central nervous system.

This work was supported by the MRC and The Wellcome Trust.

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