Effect and Mechanism of Cerebrovasodilation Induced by Total Flavonoids of Chuzhou Chrysanthemum in Rats

To investigate cerebral vasorelaxation of total flavonoids of Chuzhou chrysanthemum (TFCC) in rats and its mechanism.

Cerebral basilar arteries (CBA) of rats were isolated, and the vasodilation induced by TFCC (10-2,560 mg/L) following pretension with 100 nmol/L U₄₆₆₁₉ or 30 mmol/L KCl were measured using a pressure myograph system. Addition of H₂S synthase cystathionine-γ-lyase (CSE) inhibitor dl-propargylglycine (PPG, 100 µ mol/L), a large-conductance Ca²⁺-activated potassium (BK_Ca) channel blocker iberiotoxin (IBTX,100 nmol/L) and L-type Ca²⁺ channel blocker nifedipine (100 nmol/L) were added to determine the effect of pretreatment with the inhibitors on TFCC-induced vasorelaxation. KCl (30 mmol/L) was used as a contractile agent, TFCC (3.3-270 mg/L)-induced relaxation was detected by measuring the length of the long axis of rat cerebral vascular smooth muscle cells (VSMCs). Determination of the effect of pretreatment of VSMCs by IBTX or nifedipine on TFCC-induced cellular relaxation, and intracellular free Ca²⁺ concentration ([Ca²⁺]_i) was detected by fluorescent method. Endothelial cells (ECs) were co-cultured with VSMCs to observe the effect of endogenous H₂S on TFCC-induced relaxation in VSMCs.

TFCC caused a concentration-dependent vasorelaxation in the rat CBA precontracted with KCl or U₄₆₆₁₉ (P<0.01). Endothelial removal markedly attenuated this vasodilation, but the remaining vasorelaxation was still significant (P<0.01). The TFCC-induced cerebral vasorelaxation was remarkably inhibited by PPG, IBTX and nifedipine (P<0.01). TFCC caused a significant relaxation of rat CBA VSMCs (P<0.01). Co-culture with wild-type cerebral ECs but not with cystathionine-γ-lyase- or 3-mercaptosulfotransferase-knockout ECs markedly enhanced TFCC-induced relaxation of VSMCs (P<0.05) and increased H₂S content (P<0.01). TFCC decreased the [Ca²⁺]_i in VSMCs (P<0.01), which was attenuated by PPG and IBTX.

TFCC dilated rat CBA in both endothelium-dependent and -independent manner. Its endothelium-dependent dilation was probably involved in the blockade of L-type Ca²⁺ channels caused by endothelial H₂S activating BK_Ca channels in VSMCs; its endothelium-independent relaxation was primarily from the direct blockade of the L-type Ca²⁺ channels in VSMCs.