Calcium Channel Blockade Ameliorates Endoplasmic Reticulum Stress in the Hippocampus Induced by Amyloidopathy in the Entorhinal Cortex

Abstract

Entorhinal cortex (EC) is one of the first cerebral regions affected in Alzheimer's disease (AD). The pathology propagates to neighboring cerebral regions through a prion-like mechanism. In AD, intracellular calcium dyshomeostasis is associated with endoplasmic reticulum (ER) stress. This study was designed to examine hippocampal ER stress following EC amyloidopathy. Aβ1-42 was bilaterally microinjected into the EC under stereotaxic surgery. Rats were daily treated with 30 μg of isradipine, nimodipine, or placebo over one week. Passive avoidance and novel object recognition (NOR) tasks were performed using shuttle box and NOR test, respectively. GRP78/BiP and CHOP levels were measured in the hippocampal dentate gyrus (DG) by western blot technique. The glutathione (GSH) level and PDI activity were also assessed in the hippocampus by colorimetric spectrophotometer. Aβ treated group developed passive avoidance and novel recognition memory deficit compared to the control group. However, treatment with calcium channel blockers reversed the impairment. BiP and CHOP level increased in the hippocampus following amyloidopathy in the EC. PDI activity and GSH level in the hippocampus decreased in the Aβ treated group, but calcium channel blockers restored them toward the control level. In conclusion, memory impairment due to EC amyloidopathy is associated with ER stress related bio-molecular changes in the hippocampus, and treatment with L-type calcium channel blockers may prevent the changes and ultimately improve cognitive performance.

Keywords: Alzheimer disease; Calcium channel blockers; Endoplasmic reticulum stress; Entorhinal cortex; Protein disulfide-isomerases.

Figure 1

Calcium channel blockers attenuated the deteriorative effect of Aβ on passive avoidance learning and memory. (A) Aβ treated rats showed less latency to enter the dark compartment (step through latency) compared to the control group and treatment with isradipine or nimodipine could increase the latency toward the control level. (B) Microinjection of Aβ into the entorhinal cortex increased the time spent in the dark compartment in contrast to the control group, and treatment by isradipine or nimodipine reverse the deteriorative effect of Aβ. Data are expressed as mean ± SEM. **p < 0.01 compared to the control and #p < 0.05, ##p < 0.01 compared with the Aβ group

Figure 2

Calcium channel blockers restore the novel object recognition performance in AD model of rats. (A) Control group spent more time to explore the novel object (NOE) compared to the familiar object. (B) However, no difference of exploration time between familiar and novel object was observed in Aβ treated rats. (C) and (D) Treatment of AD rats with isradipine or nimodipine could restore the NOR task. Data are expressed as mean ± SEM

Figure 3

Amyloidopathy in the entorhinal cortex induced alteration of ER stress associated biomolecules in the hippocampal dentate gyrus. (A) The representative blots of BiP and CHOP obtained by western blot. (B) Microinjection of Aβ into the entorhinal cortex resulted in higher BiP level in the dentate gyrus. However, daily microinjection of isradipine restored it to the control level. (C) CHOP level in the hippocampus was increased following amyloidopathy in the entorhinal cortex and treatment with isradipine could prevent its rise in the DG of Aβ treated rats. Data are expressed as mean ± SEM. ^*p < 0.05, ^**p < 0.01 and ^***p < 0.001 compared with the Aβ treated group

Figure 4

Decreased PDI activity in the hippocampus following microinjection of Aβ into the entorhinal cortex was reversed by calcium channel blockers, isradipine or nimodipine. The PDI-catalyzed reduction of insulin in the presence of DTT was measured to assay PDI activity (n = 4). ^**p < 0.01 compared to the control group and ^##p < 0.01, ^###p < 0.001 compared to the Aβ group

Figure 5

Effect of intra-EC microinjection of Aβ on GSH level in the hippocampus. Microinjection of Aβ into the EC decreased the GSH level in the hippocampus compared to the control group. However, treatment with isradipine or nimodipine could restore the GSH level in the Aβ treated group (n = 4-5 in all group). (^*p < 0.05, ^**p < 0.01 and ^***p < 0.001 compared with the Aβ