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, 20 (1), 149-55

Neuronal-glial Interactions Mediated by interleukin-1 Enhance Neuronal Acetylcholinesterase Activity and mRNA Expression

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Neuronal-glial Interactions Mediated by interleukin-1 Enhance Neuronal Acetylcholinesterase Activity and mRNA Expression

Y Li et al. J Neurosci.

Abstract

Cholinergic dysfunction in Alzheimer's disease has been attributed to stress-induced increases in acetylcholinesterase (AChE) activity. Interleukin-1 (IL-1) is overexpressed in Alzheimer's disease, and stress-related changes in long-term potentiation, an ACh-related cerebral function, are triggered by interleukin-1. Microglial cultures (N9) synthesized and released IL-1 in response to conditioned media obtained from glutamate-treated primary neuron cultures or PC12 cells. This conditioned media contained elevated levels of secreted beta-amyloid precursor protein (sAPP). Naive PC12 cells cocultured with stimulated N9 cultures showed increased AChE activity and mRNA expression. These effects on AChE expression and activity could be blocked by either preincubating the glutamate-treated PC12 supernatants with anti-sAPP antibodies or preincubating naive PC12 cells with IL-1 receptor antagonist. These findings were confirmed in vivo; IL-1-containing pellets implanted into rat cortex also increased AChE mRNA levels. Neuronal stress in Alzheimer's disease may induce increases in AChE expression and activity through a molecular cascade that is mediated by sAPP-induced microglial activation and consequent overexpression of IL-1.

Figures

Fig. 1.
Fig. 1.
IL-1β induction of AChE mRNA in vivo. Illustration (A) and quantification (B) of AChE and G3PDH mRNA levels in cortex of rats after 21 d exposure to slow-release pellets containing IL-1β or vehicle (sham) or unoperated normal rat. Values are expressed as mean ± SEM. **p < 0.01, significantly different from control.
Fig. 2.
Fig. 2.
Induction of acetylcholinesterase activity in primary neuronal cultures after treatment with IL-1β (100 ng/ml for 24 hr). Cultures were derived from cerebral cortex (Ctx), basal forebrain (Bfb), hippocampus (Hi), or cerebellum (Cb). Values are expressed as mean ± SEM for four replicates. **p < 0.01, significantly different from control.
Fig. 3.
Fig. 3.
Phase-contrast photomicrographs of treated and untreated, differentiated and undifferentiated PC12 cell cultures (A–D) and of primary cortical neuronal cultures (E, F). A, Undifferentiated, untreated PC12 cell cultures. B, Undifferentiated PC12 cell cultures treated with IL-1β (100 ng/ml for 24 hr). C, Differentiated (NGF-induced), untreated PC12 cell cultures. D, Differentiated PC12 cell cultures treated with IL-1β (100 ng/ml for 24 hr). There are no discernible morphological difference between IL-1β-treated and untreated PC12 cell cultures. E, F, Acetylcholinesterase histochemical reaction of primary cortical neuronal cultures in the absence (E) or presence (F) of IL-1β (100 ng/ml for 24 hr). There was no observable difference in the numbers of AChE-positive cells in response to IL-1β.A–D are the same magnification, and Eand F are the same magnification. Scale bars, 60 μm.
Fig. 4.
Fig. 4.
IL-1β induction of AChE activity in PC12 cell cultures. A, Dose-dependent induction of AChE activity after treatment with IL-1β, IL-1β plus IL-1ra, TNFα alone, or IL-6 alone. B, Time course of the IL-1β (100 ng/ml) effect on undifferentiated PC12 cell cultures. Values expressed as mean ± SEM for five replicates. *p < 0.05, **p < 0.01, significantly different from corresponding control values.
Fig. 5.
Fig. 5.
IL-1β induction of AChE activity in NGF-differentiated PC12 cell cultures. Values expressed as mean ± SEM for eight replicates. **p < 0.001, significantly different from corresponding control values.
Fig. 6.
Fig. 6.
Induction of AChE mRNA expression in PC12 cell cultures after treatment with IL-1β (100 ng/ml for 24 hr). Values are expressed as mean ± SEM for four replicates. **p < 0.01, significantly different from control.
Fig. 7.
Fig. 7.
Induction of AChE protein expression in PC12 cell cultures after treatment with IL-1β (100 ng/ml for 24 hr). Values are expressed as mean ± SEM for six replicates. *p = 0.034, significantly different from control.
Fig. 8.
Fig. 8.
Illustration of increased βAPP synthesis in (A) and release by (B) PC12 cell cultures, after 30 min of glutamate (1 mm) treatment. Illustration by RT-PCR of expression of IL-1β mRNA levels in N9 cell lysates (C) from N9 cells that were not exposed to medium from PC12 cell (Con) or were treated with medium from untreated PC 12 cells (Unt), medium from glutamate-treated PC12 cells (Glut), or medium from glutamate-treated PC12 cells preabsorbed with anti-sAPP antibody (Glut+anti-sAPP). Illustration of IL-1β in N9 cell media by immunoprecipitation (D) after treatment of N9 cell cultures with either medium from untreated PC 12 cells (Unt) or medium from glutamate-treated PC12 cells (Glut).
Fig. 9.
Fig. 9.
AChE activity in PC12 cells cocultured with N9 cells. Before coculture with naive PC12 cells, N9 cells either were not preincubated with PC12 cell medium (Con) or were preincubated with medium from naive PC12 cells (Unt), media from glutamate-treated PC12 cells (Glut), or medium from glutamate-treated PC12 cells preabsorbed with anti-sAPP antibody (Glut/anti-sAPP). Before coculture with N9 cells that were preincubated with glutamate-treated medium, naive PC12 cells were pretreated with IL-1 receptor antagonist (Glut/IL-1ra). Values are expressed as mean ± SEM for at least five replicates. **p < 0.01, significantly different from control.

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