Combination therapy with A1 receptor agonist and vitamin C improved working memory in a mouse model of global ischemia-reperfusion

Abstract

Introduction: Stroke is one of the most important reasons of death. Hence, trials to prevent or lessen the complications originated by stroke are a goal of public health worldwide. The ischemia-reperfusion causes hypoxia, hypoglycemia and incomplete repel of metabolic waste products and leads to accumulation of free radicals triggering neuronal death. The A1 adenosine receptoras an endogenous ligand of adenosine is known to improve cell resistance to destructive agentsby preventing apoptosis. Vitamin C as a cellular antioxidant is also known as an effective factor to reduce damages initiated by free radicals. We studied the protective effects of A1 receptor agonist in combination with vitamin C against ischemia-reperfusion.

Methods: Ischemia was induced by common carotid artery occlusion in bulb-c mice (20-30 gr). Y-Maze was employed to scale the short-term memory and Nissl staining was used to count the cells in hippocampus.

Results: We found that concurrent treatment of A1 receptor agonist and vitamin C significantly reduced neuronal death in CA1. The Memory scores were also significantly improved (P < 0.05).

Discussion: Our data point to the therapeutic effects of CPA/vitamin C co-administration and highlight the beneficial role of A1 adenosine receptor signaling in the context of stroke.

Keywords: A1 receptor; Hippocampus; Ischemia-Reperfusion; Vitamin C.

Figure 1

Comparison of short term memory status among the experimental groups Short term memory status was measured using Y maze. Values are shown as mean ± SEM. Ischemic group represents significant difference compare to the intact group (*P < 0.05). Treatment groups, except DMSO and DPCPX treated groups represent significant difference compared to ischemic group (*P < 0.05). The group named CPA/AA treated with CPA and vitamin C showed a significant increase in Y-Maze results, compared to the CPA or vitamin C groups (±P < 0.05).

Figure 2

Neuronal death in CA1 region of hippocampus Cresyl violet staining of brain sections from experimental groups was performed at the end of treatment used to evaluate the neuronal density and structure. The Animals treated with CPA (f) or vitamin C (g) had more cell density which is representative of less dead neurons compared to the ischemia group (b). Co-administration of both CPA and vitamin C reduces cell death (h) compared to ischemia groups and groups received CPA (f) or vitamin C (g). Administration of A1 receptor antagonist (DPCPX) intensified the cell death among ischemic neurons and reduced cell density (d). White and black arrows are representative of normal and dead cells, respectively. Experimental groups including Intact, Ischemia, DMSO, DPCPX, DPCPX/AA, CPA, AA (vitamin C) and CPA/AA are shown a, b, c, d, e, f, g, h digital images prepared at 40 x magnifications, respectively. Scale bars: 200 µm.

Figure 3

Comparison of the normal cells density in the CA1 region of hippocampus Values are shown as mean ± SEM. Cell density was measured by counting normal neurons in CA1 region using cresyl violet staining. Cell density in groups treated with CPA or vitamin C (shown with AA) is significantly increased compared to the ischemic group (*P < 0.05). The group named CPA/ AA, treated with both CPA and vitamin C showed a significant increase in density of normal cells in the CA1 region compared to the CPA or vitamin C (AA) groups (±P < 0.05).