Effects of Tetramethylpyrazine on Functional Recovery and Neuronal Dendritic Plasticity after Experimental Stroke

Abstract

The 2,3,5,6-tetramethylpyrazine (TMP) has been widely used in the treatment of ischemic stroke by Chinese doctors. Here, we report the effects of TMP on functional recovery and dendritic plasticity after ischemic stroke. A classical model of middle cerebral artery occlusion (MCAO) was established in this study. The rats were assigned into 3 groups: sham group (sham operated rats treated with saline), model group (MCAO rats treated with saline) and TMP group (MCAO rats treated with 20 mg/kg/d TMP). The neurological function test of animals was evaluated using the modified neurological severity score (mNSS) at 3 d, 7 d, and 14 d after MCAO. Animals were euthanized for immunohistochemical labeling to measure MAP-2 levels in the peri-infarct area. Golgi-Cox staining was performed to test effect of TMP on dendritic plasticity at 14 d after MCAO. TMP significantly improved neurological function at 7 d and 14 d after ischemia, increased MAP-2 level at 14 d after ischemia, and enhanced spine density of basilar dendrites. TMP failed to affect the spine density of apical dendrites and the total dendritic length. Data analyses indicate that there was significant negative correlation between mNSS and plasticity measured at 14 d after MCAO. Thus, enhanced dendritic plasticity contributes to TMP-elicited functional recovery after ischemic stroke.

Figure 1

The structure of TMP.

Figure 2

The schematic diagram of ischemic penumbra (IP).

Figure 3

A simple flow-chart of experimental design.

Figure 4

A representative photograph of TTC staining of MCAO rat.

Figure 5

Effect of TMP on neurological status in rats with ischemic cerebral injury. The data were presented as mean ± standard deviation (n = 12). ^* P < 0.05 between model group and TMP group; ^** P < 0.01 between model group and TMP group.

Figure 6

The expression levels of MAP-2 within peri-infarct area of three groups in sham, model, and TMP groups at 3 d, 7 d, and 14 d after MCAO. (a) Immunohistochemical staining of three groups (400x). (b) MAP-2 levels of three groups through measuring the integral optical density (IOD). Data were presented as mean ± standard deviation (n = 6). ^* P < 0.01 and ^** P < 0.001.

Figure 7

A representative dendritic morphology of layer V pyramidal cells of rats (Golgi-Cox staining). Photomicrograph was viewed at ×200 magnification. Bar = 50 μm.

Figure 8

Quantification analysis of effect of TMP on total dendritic length using Sholl analysis. Data were presented as mean ± standard deviation (n = 6).

Figure 9

Quantification analyses of effect of TMP on dendritic spine density (basilar dendrites and apical dendrites, resp.). (a) The segments were acquired from layer V pyramidal cells and viewed at ×1000 magnification. Scale bar = 10 μm for all segments. (b) The dendritic spine density was expressed as spines/10 μm and the data were presented as mean ± standard deviation (n = 6). ^* P < 0.05 and ^** P < 0.01.

Figure 10

Scatterplots present correlations analysis of mNSS and plasticity measured at 14 d after MCAO. (a) Scatterplots of mNSS and MAP-2 level. (b) Scatterplots of mNSS and total dendritic length. (c) Scatterplots of mNSS and spine density of basilar dendrites. (d) Scatterplots of mNSS and spine density of apical dendrites.