Protective effect of compound Danshen (Salvia miltiorrhiza) dripping pills alone and in combination with carbamazepine on kainic acid-induced temporal lobe epilepsy and cognitive impairment in rats

Abstract

Context: Temporal lobe epilepsy (TLE) is resistant to antiepileptic drugs (AEDs) and is associated with cognitive impairment. The modern Chinese medicine, compound Danshen dripping pills (CDDP), is clinically effective in treating epilepsy and improving cognitive impairment.

Objective: This study evaluated the protective effects of CDDP alone and in combination with carbamazepine (CBZ) on kainic acid-induced TLE and cognitive impairment in rats.

Materials and methods: Sprague-Dawley rats were randomly divided into five groups: control (sham operated), model, CDDP, CBZ and combined. A TLE model was then created via bilateral intrahippocampal injection of 0.35 μg kainic acid (KA). Rats received CDDP (85 mg/kg), CBZ (100 mg/kg) or combined (85 mg/kg CDDP +100 mg/kg CBZ) via intragastric administration for 90 d, respectively. Seizure intensity, apoptosis and glial cell line-derived neurotrophic factor (GDNF) were measured. Furthermore, the improvement in cognitive impairment and hippocampal neuronal damage was evaluated.

Results: CDDP combined with CBZ significantly decreased seizure severity and frequency (p < 0.05) and ameliorated cognitive impairment (p < 0.05). The model group showed a significant reduction of neurons and Bcl-2/Bax expression in the hippocampus CA3 area (p < 0.01), the combined groups significantly reversed these change (p < 0.01). GDNF expression in the combined groups showed a clear increase over the model group (p < 0.05).

Conclusion: These findings support the use of CDDP as an adjuvant drug for the treatment of TLE and cognitive deficit. Its mechanism might be related to an anti-apoptosis effect and up-regulation of GDNF.

Keywords: Intractable epilepsy; anti-apoptosis; antiepileptic drugs; glial cell line-derived neurotrophic factor; hippocampal CA3 area; traditional Chinese medicine.

Figure 1.

An overview of the experimental design.

Figure 2.

Effects of CDDP and its combination with CBZ on the degree of seizure. (A) Seizure severity score. (B) Frequency of SRS. SRS in each group were recorded three times a day for 1 week, only the seizures of stage 3 or greater according to Racine stages were recorded. The results are presented as mean ± SEM. *p < 0.05 versus the model group (n = 14 per group).

Figure 3.

Effect of CDDP and its combination with CBZ on cognitive impairment in the MWM place navigation test. Average escape latencies to find the hidden platform for each trial day are presented as means ± SEM (n = 6 per group).

Figure 4.

Effect of CDDP and its combination with CBZ on cognitive impairment in the MWM probe trial. (A) Frequency of platform crossing. (B) Time spent in the target quadrant (%). Results are presented as means ± SEM. △p < 0.05, △△p < 0.01 versus the control group; *p < 0.05, **p < 0.01 versus the model group; ▲p < 0.05 versus the CBZ group (n = 6 per group).

Figure 5.

Effect of CDDP and its combination with CBZ on KA-induced neuron death (A) Nissl staining was used to assess the neuronal damage of hippocampus (magnification ×50). The arrowheads show the position of the hippocampal CA3 area. (B) The neuronal loss in the hippocampal CA3 region (magnification ×200). (C) The number of surviving neurons in the hippocampal CA3 region was counted. The results are presented as means ± SEM. △△p < 0.01 versus the control group; **p < 0.01 versus the model group; ▽▽p < 0.01 versus the CDDP group; ▲p < 0.05 versus the CBZ group (n = 6 per group). Scale bars: 100 μm.

Figure 6.

Effect of CDDP and its combination with CBZ on the expression of GDNF and Bcl-2/Bax in the hippocampal CA3 region. (A) Western blotting was used to evaluate protein expression of GDNF, Bcl-2 and Bax, and β-actin was used as an internal control. (B) Densitometry analysis was performed using Bio-Rad Quantity One software. The relative band density of GDNF and Bcl-2/Bax are shown in the bar diagram. The results are presented as means ± SEM. △p < 0.05 versus the control group; *p < 0.05 versus the model group; ▽p < 0.05 versus the CDDP group (n = 6 per group).