Effects of agmatine on cognitive functions during vascular dementia in biological aging through eNOS and BDNF expression

Figures & data

Table 1. Body weights (g), blood glucose (mg/dl) and systolic BP (mm/Hg), locomotor activity, passive avoidance acquisition test phases and MWMT of the time spent in the target quadrant on day 5 of rats in 4- (4 M), 18-(18 M), 24-month-old (24 M) and agmatine-treated age-matching groups.

	4 M control (n = 6)	4 M+ Agmatine (n = 6)	18 M (n = 6)	18M + Agmatine (n = 6)	24 M (n = 6)	24 M + Agmatine (n = 6)
Body weight (g)	266.83 ± 6.42	275.33 ± 4.89	311.16 ± 4.1*	312.33 ± 4.16*	441 ± 6.31*	431.83 ± 6.36*
SBP (mm/Hg)	103.9 ± 3.034	102.7 ± 3.20	109.9 ± 1.18	104.4 ± 2.94	114 ± 1.46	109.5 ± 2.21
Blood glucose (mg/dl)	88.67 ± 3.72	89.17 ± 3.33	88.16 ± 2.47	88.83 ± 3.56	84.5 ± 3.26	80.17 ± 5.56
Total locomotor activity	1135 ± 48.41	1065 ± 57.04	1031 ± 40.65	1054 ± 54.44	938.8 ± 57.86	971.3 ± 71.71
First-day latency (s)	80.95 ± 21.57	69.68 ± 14.92	80.99 ± 25.10	99.98 ± 33.12	79.92 ± 21.70	78.67 ± 25.59
Time spent in escape platform’s quadrant (s)	28.67 ± 2.36	29.42 ± 1.80	19.33 ± 0.61*	22.17 ± 1.80	15.75 ± 1.40*	23.33 ± 1.94^#

Note: Values are arithmetic means ± SEM, n = number of animals used.

*P < .05, statistically different from 4-month-old control rats and ^#P < .05, statistically different from 24-month-old rats.

Figure 1. Passive avoidance retention test phases of young (4-month-old), middle-aged (18-month-old), aged (24-month-old) and agmatine (40 mg/kg)-treated matching groups (n = 12 rats in each group). Data were presented as mean ± SEM and *P < .05 and ***P < .001 compared to the young control group (4-month-old).

Figure 2. Escape latency (s) in MWMT results of 1–4 days by rats in young (4-month-old), middle-aged (18-month-old), aged (24-month-old) and agmatine (40 mg/kg)-treated matching groups (n = 12 rats in each group). Data were presented as mean ± SEM and **P < .01 and ***P < .001 compared to the young control group (4-month-old).

Figure 3. Representative image illustrating BDNF expression (arrows) in hippocampal formation. Control (a), 24-month (b) and 24-month + agmatine (c) groups in the CA1 region, and control (d), 24-month (e) and 24-month + agmatine (f) groups in the CA3 region. control (g), 24 month (h) and 24-month + agmatine (i) groups in the amygdala. BDNF expression was decreased in 24 month in the CA1 and CA3 regions of the hippocampus, and amygdala, whereas in 24-month + agmatine rats BDNF expression was similar to those in the control group in the all regions. Scale bars: 50 μm.

Table 2. Semi-quantitative distribution of BDNF-immunoreactive neurons of the CA1 and CA3 regions of the hippocampus and amygdala and eNOSimmunoreactivity of aorta in rats.

Hippocampal CA1	Animal 1	Animal 2	Animal 3	Animal 4	Animal 5	Animal 6
4 M	2+	1+	2+	3+	2+	3+
24 M	1+	1+	2+	1+	2+	0
24 M + agmatine	2+	1+	1+	3+	2+	2+
Hippocampal CA3
4 M	2+	2+	3+	2+	2+	2+
24 M	1+	2+	1+	0	0	2+
24 M + agmatine	1+	3+	2+	2+	2+	2+
Amygdala
4 M	3+	2+	3+	2+	2+	3+
24 M	1+	1+	1+	2+	2+	1+
24 M + agmatine	1+	3+	2+	2+	3+	2+
eNOS
4 M	3+	3+	2+	2+	2+	3+
24 M	0	1+	1+	2+	1+	1+
24 M + agmatine	2+	3+	2+	2+	3+	1+

Note: The staining intensity was classified as no expression (0), very (1+), moderate (2+), strong (3+) or very strong (4+) expression. The immunopositivity was decreased in 24-month-old rats group (24M) compared to the 4-month-old control group (4M) (P < .01, Kruskal–Wallis test). In the agmatine-treated group, both BDNF and eNOS immunoreactivity were similar to that of the control group.

Table 3. E_m (% of 10⁻⁶ M phenylephrine) values for carbachol, sodium nitroprusside and papaverine, E_m values (g) for 80 mM KCl and pD₂ values (–log EC₅₀) for carbachol, and sodium nitroprusside in rings of thoracic aorta obtained from 4- (4 M), 18-(18 M), 24-month-old (24 M) and agmatine-treated age-matching group rats.

	4 M control (n = 6)	4 M + Agmatine (n = 6)	18 M (n = 6)	18 M + Agmatine (n = 6)	24 M (n = 6)	24 M + Agmatine (n = 6)
KCl Em (g)	0.70 ± 0.10	0.71 ± 0.09	0.88 ± 0.12	0.79 ± 0.07	0.77 ± 0.10	0.77 ± 0.11
Papaverine Em (%)	100 ± 0	100 ± 0	100 ± 0	100 ± 0	100 ± 0	100 ± 0
Carbachol Em (%)	61.8 ± 3.99	56.7 ± 5.52	37.5 ± 3.83*	48.5 ± 7.1	29.2 ± 3.7*	46.8 ± 3.4
Carbachol pD2	6.82 ± 0.54	6.89 ± 0.32	6.72 ± 0.36	6.82 ± 0.42	5.74 ± 0.39*	6.80 ± 0.52
SNP Em (%)	100 ± 0	100 ± 0	100 ± 0	100 ± 0	100 ± 0	100 ± 0
SNP pD2	6.38 ± 0.24	6.30 ± 0.22	6.46 ± 0.36	6.46 ± 0.32	5.92 ± 0.52	6.39 ± 0.44

Note: Values are arithmetic means ± SEM. n = the number of thoracic aortic rings used.

*Statistical significance was P < .05.

Figure 4. Carbachol concentration–response curves in isolated thoracic aortic rings pre-contracted with phenylephrine (10⁻⁶ M). The concentration–response curve for carbachol was shifted to the right with significantly lower values of E_m and pD₂ in the 24-month-old group than in 4-month-old controls (*P < .05). Impairment of relaxation in the agmatine treatment group was returned to that seen in the controls. Each point is expressed as a percentage of the contraction induced by phenylephrine and is represented as the mean ± SEM. The values in parentheses indicate the number of preparations used; *P < .05, different from the response of tissue rings from 4-month-old young control rats.

Figure 5. Sodium nitroprusside concentration–response curves in isolated thoracic aortic rings pre-contracted with phenylephrine (10⁻⁶ M). The relaxation response to SNP was similar among vascular tissues from all groups. Each point is expressed as a percentage of the contraction induced by phenylephrine and is expressed as the mean ± SEM. The values in parentheses indicate the number of preparations used.

Figure 6. Representative light microscopy of control, 24-month-old and agmatine-treated 24-month-old groups in the endothelium of the aorta. Decreased eNOS (arrows) (b) immunoreactivity in the aorta in 24-month-old rats compared to 4-month-old controls (a) and increased eNOS (c) immunoreactivity in endothelial tissue in the agmatine-treated 24-month-old group compared to 24-month-old groups (b). Scale bars: 20 μm.