5‑HT₄ Receptors Constitutively Promote the Non-Amyloidogenic Pathway of APP Cleavage and Interact with ADAM10

In addition to the amyloidogenic pathway, amyloid precursor protein (APP) can be cleaved by α-secretases, producing soluble and neuroprotective APP alpha (sAPPα) (nonamyloidogenic pathway) and thus preventing the generation of pathogenic amyloid-β. However, the mechanisms regulating APP cleavage by α-secretases remain poorly understood. Here, we showed that expression of serotonin type 4 receptors (5-HT₄Rs) constitutively (without agonist stimulation) induced APP cleavage by the α-secretase ADAM10 and the release of neuroprotective sAPPα in HEK-293 cells and cortical neurons. This effect was independent of cAMP production. Interestingly, we demonstrated that 5-HT₄ receptors physically interacted with the mature form of ADAM10. Stimulation of 5-HT₄ receptors by an agonist further increased sAPPα secretion, and this effect was mediated by cAMP/Epac signaling. These findings describe a new mechanism whereby a GPCR constitutively stimulates the cleavage of APP by α-secretase and promotes the nonamyloidogenic pathway of APP processing

Spatial learning and reference memory of the 5-HTR₄ knock-out mice (experiment 1).

<p>Left-hand side (A): Performance over days (2–14) for the acquisition (days 2–10) and the reversal (days 11–14) of spatial memory testing in the Morris water maze for wild-type (WT) and 5-HTR₄ knock-out (KO) mice. The score on each day represents the mean ± s.e. of path length (cm). Right-hand side (B): Probe trials (60 s) on days 5, 10, 14, 15, and 20. Mean ± s.e. path length in the target platform zone/sum of path lengths in the four equivalent zones (target PF/totPFs) in WT and KO mice. Horizontal lines represent chance level.</p

Spatial learning and reference memory of the 5-HTR₄ knock-out mice (experiment 2).

<p>Effect of scopolamine injection (0.8 mg/kg, i.p., 20 min) on performance. Left-hand side (A): mean ± s.e. of path length (cm) over days (1–4) in wild type (WT) and 5-HTR₄ knock-out (KO) mice. Right-hand side (B): Probe trials (60 s) on days 4 and 5. Top: Mean ± s.e. path length in the whole pool (swim speed) of the WT and KO mice. Bottom: Mean ± s.e. path length in the target platform zone/sum of path lengths in the four equivalent zones (target PF/totPFs) in WT and KO mice. Horizontal lines represent chance level. A significant treatment effect is noted (* p = 0.05, ** p = 0.02).</p

Reduced enzymatic activity of ChAT in the absence of 5-HTR₄.

<p>Values are means ± s.e.m. of ChAT activity expressed in <i>p</i>mol ACh/min/µg protein for 7–8 WT and 6–7 KO mice in baseline conditions and 3–4 WT and 3–4 KO mice following the session of behavioral tests. A significant difference between genotypes or conditions is marked (* p<0.05, ** p<0.01 and §§ p<0.01, §§§ p<0.001, respectively). The genotype x condition interaction is significant (# p<0.05).</p

The effects of scopolamine on the enzymatic activity of ChAT were suppressed in the absence of 5-HTR₄.

<p>Values are means ± s.e.m. of Ach levels expressed in pmol/min/µg protein for saline-treated mice (n = 3−4 WT, n = 3−4 KO) and for scopolamine-treated mice (n = 4−5 WT, n = 4−5 KO) following the behavioral test session. A significant difference between genotypes or treatments is noted (* p<0.05 and § p<0.05, §§ p<0.01, respectively). The genotype x treatment interaction is significant (# p<0.05, ## p<0.01).</p