Permeability of dopamine D2 receptor agonist hordenine across the intestinal and blood-brain barrier in vitro

Abstract

Hordenine, a bioactive food compound, has several pharmacological properties and has recently been identified as a dopamine D2 receptor (D2R) agonist. Since the pharmacokinetic profile of hordenine has been described to a limited extent, the present study focused on the transfer and transport of hordenine across the intestinal epithelium and the blood-brain barrier (BBB) in vitro. Hordenine was quickly transferred through the Caco-2 monolayer in only a few hours, indicating a rapid oral uptake. However, the high bioavailability may be reduced by the observed efflux transport of hordenine from the bloodstream back into the intestinal lumen and by first pass metabolism in intestinal epithelial cells. To determine the biotransformation rate of hordenine, the metabolite hordenine sulfate was synthesized as reference standard for analytical purposes. In addition, transfer studies using primary porcine brain capillary endothelial cells (PBCEC) showed that hordenine is able to rapidly penetrate the BBB and potentially accumulate in the brain. Thus, a D2R interaction of hordenine and activation of dopaminergic signaling is conceivable, assuming that the intestinal barrier can be circumvented by a route of administration alternative to oral uptake.

Fig 1

Chemical structures of hordenine (A) and its metabolite hordenine sulfate (B).

Fig 2

Extracted ion chromatograms of standard solution containing 500 nM hordenine and hordenine sulfate diluted in MeOH/H₂O (1+19, v/v) (A) and in cell culture matrix (B). Both qualifier and quantifier MRM transitions are shown. The matrix signal (intensity: 1,52×10⁵ cps) is resulting from cell culture medium.

Fig 3. Relative transepithelial/transendothelial electrical resistance (TEER) and electrical capacitance (c_CL,) of Caco-2 cells (A1/A2) and primary porcine brain capillary endothelial cells (PBCEC, B1/B2) incubated with 1 μM hordenine or solvent (neg, 0.1% DMSO) for 48 h (n = 3 × 3).

The data are normalized to the initial values at the beginning of the transport studies and represented as means ± standard deviations. Significant differences (p < 0.05) between the test substances and the respective negative control were calculated in the Student’s t-test and marked with asterisks.

Fig 4

Passive transfer (A) and active transport (B) of hordenine through the Caco-2 monolayer over the course of 48 h. The passive transfer of 1 μM hordenine from apical (ap) to basolateral (ba) compartment was determined (n = 3 × 3). Hordenine and hordenine sulfate (abbreviated as sulfate) were quantified using HPLC-MS/MS and normalized to the initial amount (0.76 nmol) on the apical side. For active transport experiments equimolar concentrations of 200 nM hordenine were applied in the apical and basolateral compartment (n = 3 × 3). Data are presented as means ± standard deviations. Statistically significant differences between concentration in the two compartments are marked by asterisks (*: p ≤ 0.05; **: p ≤ 0.01; ***: p ≤ 0.001).

Fig 5

Passive transfer (A) and active transport (B) of hordenine through the PBCEC monolayer. The passive transfer of hordenine from apical (ap) to basolateral (ba) compartment was determined after apical application of 1 μM hordenine (n = 3 × 3). Hordenine was quantified using HPLC-MS/MS and normalized to the initial amount (0.76 nmol) on the apical side. The active transport experiments based on the recovery of 200 nM hordenine in the apical and basolateral compartment after incubation of equimolar concentrations in both compartments (n = 3 × 3). Data are presented as means ± standard deviations.

Fig 6

Schematic illustration of transport and metabolism of hordenine through the intestinal barrier (A) and the blood-brain barrier (B).