Gintonin facilitates brain delivery of donepezil, a therapeutic drug for Alzheimer disease, through lysophosphatidic acid 1/3 and vascular endothelial growth factor receptors

Abstract

Background: Gintonin is a ginseng-derived exogenous G-protein-coupled lysophosphatidic acid (LPA) receptor ligand, which exhibits in vitro and in vivo functions against Alzheimer disease (AD) through lysophosphatidic acid 1/3 receptors. A recent study demonstrated that systemic treatment with gintonin enhances paracellular permeability of the blood-brain barrier (BBB) through the LPA1/3 receptor. However, little is known about whether gintonin can enhance brain delivery of donepezil (DPZ) (Aricept), which is a representative cognition-improving drug used in AD clinics. In the present study, we examined whether systemic administration of gintonin can stimulate brain delivery of DPZ.

Methods: We administered gintonin and DPZ alone or coadministered gintonin with DPZ intravenously or orally to rats. Then we collected the cerebral spinal fluid (CSF) and serum and determined the DPZ concentration through liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis.

Results: Intravenous, but not oral, coadministration of gintonin with DPZ increased the CSF concentration of DPZ in a concentration- and time-dependent manner. Gintonin-mediated enhancement of brain delivery of DPZ was blocked by Ki16425, a LPA1/3 receptor antagonist. Coadministration of vascular endothelial growth factor (VEGF) + gintonin with DPZ similarly increased CSF DPZ concentration. However, gintonin-mediated enhancement of brain delivery of DPZ was blocked by axitinip, a VEGF receptor antagonist. Mannitol, a BBB disrupting agent that increases the BBB permeability, enhanced gintonin-mediated enhancement of brain delivery of DPZ.

Conclusions: We found that intravenous, but not oral, coadministration of gintonin facilitates brain delivery of DPZ from plasma via LPA1/3 and VEGF receptors. Gintonin is a potential candidate as a ginseng-derived novel agent for the brain delivery of DPZ for treatment of patients with AD.

Keywords: Alzheimer disease; Brain delivery; Donepezil; Ginseng; Gintonin.

Fig. 1

Schematic diagram of gintonin (GT)-mediated enhancement of delivery of donepezil (DPZ) in the brain. CSF, cerebrospinal fluid; IP, intraperitoneal; IV, intravenous.

Fig. 2

Effects of gintonin on circulating LPA levels in rats. (A–D) Vehicle or gintonin (40 mg/kg) was administered intravenously into catheterized rats into the tail vein, and plasma levels of various LPAs were determined at the indicated time points. Values depict the mean ± standard error of mean (S.E.M.) (n = 4). Experimental procedures for the measurement of LPAs were described in Materials and methods. LPA, lysophosphatidic acid.

Fig. 3

Determination of mean CSF and plasma concentrations of donepezil for estimation of brain delivery effects of gintonin. Rats were administered 3 mg/kg (A, DPZ concentration in CSF; B, DPZ concentration in serum), 4 mg/kg (C, DPZ concentration in CSF; D, DPZ concentration in serum), and 6 mg/kg (E, DPZ concentration in CSF; F, DPZ concentration in serum) donepezil alone or were coadministered with gintonin (15, 30, or 40 mg/kg) intravenously via the tail vein. CSF and plasma samples were collected within 5 min after each injection. The analysis of donepezil was described in Materials and methods. Values depict the mean ± S.E.M. (n = 10). *P < 0.05, **P < 0.01, ***P < 0.005, compared to with donepezil alone. CSF, cerebrospinal fluid.

Fig. 4

Determination of mean CSF and plasma concentrations time profile of donepezil on brain delivery effect of gintonin. CSF (A, DPZ concentration in CSF) and plasma (B, DPZ concentration in serum) samples were collected within 5, 15, and 30 min after intravenous injection of DPZ alone (6 mg/kg) or cotreatment with gintonin (30 mg/kg). Values depict the mean ± standard error of mean (S.E.M.) (n = 8). *p < 0.05, **p < 0.01, compared with donepezil alone. CSF, cerebrospinal fluid; DPZ, donepezil.

Fig. 5

Effects of pretreatment of LPA1/3 receptor (Ki16425), VEGF, VEGF antagonist (Axitinib), or mannitol on gintonin-mediated enhancement of brain delivery of donepezil. (A–B) After Ki16425 (30 mg/kg) or axitinib (25 mg/kg) was pretreated for 30 min by intraperitoneal injection, rats received administration of DPZ (6 mg/kg) or coadministration with gintonin (30 mg/kg) intravenously via the tail vein. ^#p < 0.01, compared with donepezil alone. *p < 0.01, **p < 0.001, compared with gintonin + donepezil. (C–D) Rats were treated with DPZ (6 mg/kg) + VEGF (200 μg/kg) or DPZ + VEGF + gintonin (30 mg/kg) via intravenous injection. ^#p < 0.01, compared with donepezil alone; ^##p < 0.01, compared with donepezil + gintonin. *p < 0.01, compared with gintonin + donepezil. (E–F) After mannitol (5 mg/kg; 20% in 0.9% saline) + DPZ (6 mg/kg) or mannitol + DPZ + gintonin (30 mg/kg) treatment to the rats, CSF and plasma samples were collected within 5 min. ^#p < 0.01, compared with donepezil alone; *p < 0.01, compared with mannitol + donepezil. Values depict the mean ± standard error of mean (S.E.M.) (n = 8). CSF, cerebrospinal fluid; DPZ, donepezil; VEGF, vascular endothelial growth factor.

Fig. 6

Effects of orally administered gintonin on donepezil brain delivery system. (A) Donepezil (10 mg/kg) was administered intravenously 60 min after oral treatment of gintonin (100 mg/kg). (C) Gintonin (40 mg/kg) was administered intravenously 60 min after oral treatment of donepezil (10 mg/kg). CSF (A and C) and plasma samples (B and D) were collected within 5 min. The analysis of donepezil was described in Materials and methods. Values depict the mean ± standard error of mean (S.E.M.) *p < 0.05, compared with donepezil alone (n = 8).

Fig. 7

Schematic illustration of gintonin-mediated stimulations of donepezil permeability into brain. Gintonin-mediated stimulations of donepezil permeability into brain might include two signaling pathways. One possibility is that activation of LPA1/3 receptors by gintonin induces morphological changes of brain microvessel endothelial cells through Gα_12/13-Rho-associated kinase morphological changes signaling pathway that is coupled to increases in brain permeability to donepezil, as shown in a previous report [14]. The other is through the VEGF. The activations of LPA1/3 receptors by gintonin is coupled to an increase in VEGF synthesis and release from brain microvessel endothelial cells through the Gα_q/11-phospholipase C (PLC)-Ca²⁺ signaling pathway. Released VEGFs increase brain permeability, as indicated in previous reports [18,19]. VEGF, vascular endothelial growth factor.