Huperzine A reverses cholinergic and monoaminergic dysfunction induced by bilateral nucleus basalis magnocellularis injection of beta-amyloid peptide (1-40) in rats

Abstract

(1) Huperzine A, a promising therapeutic agent for Alzheimer's disease (AD), was tested for its effects on cholinergic and monoaminergic dysfunction induced by injecting beta-amyloid peptide-(1-40) into nucleus basalis magnocellularis of the rat. (2) Bilateral injection of 10 microg beta-amyloid peptide-(1-40) into nucleus basalis magnocellularis produced local deposits of amyloid plaque and functional abnormalities detected by microdialysis. In medial prefrontal cortex, reductions in the basal levels and stimulated release of acetylcholine, dopamine, norepinephrine, and 5-hydroxytryptamine were observed. However, oral huperzine A (0.18 mg/kg, once daily for 21 consecutive days) markedly reduced morphologic abnormalities at the injection site in rats infused with beta-amyloid peptide-(1-40). Likewise, this treatment ameliorated the beta-amyloid peptide-(1-40)-induced deficits in extracellular acetylcholine, dopamine, and norepinephrine (though not 5-hydroxytryptamine) in medial prefrontal cortex, and lessened the reduction in nicotine or methoctramine-stimulated release of acetylcholine and K(+)-evoked releases of acetylcholine and dopamine. (3) The present results provide the first direct evidence that huperzine A acts to oppose neurotoxic effects of beta-amyloid peptide on cholinergic, dopaminergic, and noradrenergic systems of the rat forebrain.

Fig. 1

The injection site in bilateral NBM was verified by visual inspection. (A) Freezing slice of rat brain. Straight arrows indicate the injection site and curving arrows indicate the injection tract. (B) Atlas of rat brain at coordinate −1.8 mm from Bregma (Paxinos and Watson 1997). NBM was shown by dashed circle. (C) The overlay of freezing slice with atlas of rat brain

Fig. 2

Huperzine A enhances clearance of β-amyloid at NBM injection site. Brains were harvested 21 days after bilateral NBM injection of 10 μg Aβ_1–40 followed by oral treatment with saline (A) or huperzine A (0.18 mg/kg, B) once per day for 21 consecutive days. Representative sections are shown. Arrow indicates persistent amyloid immunoreactivity in saline-treated specimen. Scale bar = 50 μm

Fig. 3

HE-staining reveals morphologic changes in NBM after injection of 10 μg Aβ_1–40. Rats were killed 21 days after bilateral NBM injection of (A) vehicle, (B) Aβ_1–40 with oral administration of saline, (C) Aβ_1–40 with oral administration of 0.18 mg/kg huperzine A once per day for 21 consecutive days. Sections from three rats in each group were examined. Scale bar = 50 μm. Note pyknotic cells and decreased neuronal abundance in B, as compared with A and C

Fig. 4

Huperazine A and Aβ-induced deficits of mPFC ACh (A), DA (B), and NE (C). Rats were examined 21 days after bilateral NBM injection of 10 μg Aβ_1–40. Huperzine A was orally administered once per day for 21 consecutive days. Data represent mean ± S.E.M., n = 8–10 rats. *P < 0.05, **P < 0.01 versus vehicle; #P < 0.05 versus Aβ_1–40 injection

Fig. 5

Effects of Aβ and huperzine A on drug-stimulated ACh release in mPFC. On days 19, 20, or 21 after bilateral NBM injection of 10 μg Aβ_1–40, rats underwent microdialysis with nicotine (A, 5 mM), methoctramine (B, 0.2 mM) or K⁺ (C, 100 mM). These stimulations lasted 30 min, 20 min, and 30 min, respectively (arrows). Huperzine A (0.18 mg/kg) was orally administered once per day for 19 or 21 consecutive days. Results are expressed as percentage increases above baseline (averaged from three consecutive values obtained before stimulation). Mean values ± S.E.M are shown (n = 4–5 rats). *P < 0.05, **P < 0.01 versus baseline; ^a P < 0.05 versus vehicle-injected group, ^b P < 0.05 versus Aβ_1–40-injected group

Fig. 6

Effects of Aβ and huperzine A on K⁺-induced release of DA and NE from mPFC. On day zero, rats received bilateral NBM injections of 10 μg Aβ_1–40. Huperzine A (0.18 mg/kg) or saline was then administered orally once per day for 21 consecutive days. On day 21, K⁺ (100 mM) was perfused through the microdialysis probe for 30 min (arrow). Release of DA (A) and of NE (B) were expressed as percentage increases above baseline (average of three untreated points), shown as mean ± S.E.M. (n = 4–5 rats). *P < 0.05 versus baseline; ^a P < 0.05 versus vehicle-injected group, ^b P < 0.05 versus Aβ_1–40-injected group