Blood-brain barrier transport studies of organic guanidino cations using an in situ brain perfusion technique.

Blood-brain barrier (BBB) transport of essential polar substrates is mediated by specific, carrier-mediated transport proteins. The BBB transport mechanisms for polar compounds with terminal guanidino functional groups (R-NHC(NH)NH(2)) are not well defined. The goal of the present work was to investigate the BBB transport mechanism(s) for terminal guanidino substrates using an in situ brain perfusion technique. Brain region radiotracer influx clearance (Cl(in)) was calculated for representative guanidino substrates, [14C]L-arginine, [14C]aminoguanidine and [14C]guanidine, in the presence or absence of excess terminal guanidino analogues. The Cl(in) for [14C]L-arginine (0.21+/-0.0094 cm(3)/min/g wet brain weight, mean+/-S.E.M., n=four rats) was significantly decreased by 1000x concentrations of unlabeled L-arginine, N(G)-methyl-L-arginine, N(G)-,N(G)-dimethyl-L-arginine and N(G)-amino-L-arginine by approximately 83% (P<0.01; n=4-5), whereas 1000x concentrations of nitro-L-arginine, aminoguanidine and guanidine were without effect. In contrast, the respective Cl(in) of [14C]aminoguanidine and [14C]guanidine (0.0085+/-0.00039 and 0.015+/-0.0015 cm(3)/min/g, n=4, respectively) were not significantly decreased by 1000x concentrations of unlabeled aminoguanidine or guanidine. The Cl(in) values for all [14C]guanidino probes were significantly greater (P<0. 05) from that of [3H]inulin, a marker of cerebrovascular blood volume. These data suggest that the hydrophilic guanidino cations aminoguanidine and guanidine penetrate the BBB by a minor diffusional process with no appreciable transport via saturable processes. In contrast, BBB penetration of L-arginine occurs via the saturable basic amino acid transporter that has specificity for amino acid analogues possessing cationic terminal guanidino groups.

Cellular transport processes of aminoguanidine, a nitric oxide synthase inhibitor, in the opossum kidney cell culture line.

Aminoguanidine has potential pharmacologic utility for diabetes and nitric oxide - mediated inflammation. Because aminoguanidine is positively charged at physiologic pH (pK(a) approximately 10), it is unlikely that simple diffusion is a predominant mechanism for cellular penetration. This study sought to determine the transport processes by which aminoguanidine, a cationic compound, traverses across cellular membranes. In cultured opossum kidney (OK) cell monolayers, aminoguanidine transport involved both saturable and non-saturable diffusion processes. At passage numbers below 67, the observed V(max) and K(m) for saturable influx were significantly lower than that observed at passages greater than 79 (V(max): low passage, 21.2+/-7.8 pmol/(min*mg protein), n=3; versus high passage, 129.7+/-24.3 pmol/(min*mg protein), n=3, P<0.05; K(m): low passage, 23.7+/-10.8 microM, n=3; versus high passage, 101.7+/-5.6 microM, n=3, P<0.05; mean+/-S.E.M.). Nonsaturable processes were not statistically different (k(ns): low passage, 1.6+/-0.1 pmol/(min*mg protein*microM), n=3; high passage, 1.1+/-0.2 pmol/(min*mg protein*microM) n=3). Saturable influx was temperature dependent, and independent of ATP energy, sodium gradients or changes in membrane potential. Other organic cations competitively inhibited and trans-stimulated saturable influx. Aminoguanidine influx was increased in the presence of an outwardly-directed proton gradient and was inhibited in the presence of an inwardly-directed proton gradient. Correspondingly, aminoguanidine efflux was trans79) express a saturable, bi-directional carrier-mediated process to transport aminoguanidine across cellular membranes.

Theoretical pharmacokinetic and pharmacodynamic simulations of drug delivery mediated by blood--brain barrier transporters.

Pharmacokinetic/pharmacodynamic simulations were performed to assess the feasibility of central nervous system (CNS) drug delivery via endogenous transporters resident at the blood-brain barrier (BBB). Pharmacokinetic models were derived for intravenous bolus dosing of a hypothetical drug in the absence or presence of an endogenous, competing transport inhibitor. These models were linked to CNS pharmacodynamic models where the effect sites were either cell surface receptors or intracellularly localized enzymes. The response of the dependent parameter, the duration of effect (t(dur)), was examined in relationship to changes in the independent parameters, i.e. dose, elimination rate constant (k(e1)), BBB transport parameters (K(m1) and V(max1)) and EC(50) (effective concentration that elicits a 50% response). As expected, t(dur) increased with (a) increases in drug doses, (b) decreases in k(e1) or (c) decreases in EC(50), irrespective of the effect site. Surprisingly, endogenous transport inhibition produced decreases in drug terminal half-life and corresponding decreases in t(dur). Interestingly, t(dur) was independent of assigned transporter K(m) and V(max) when the dose/EC(50) ratio (dose/EC(50)) was >1 (irrespective of endogenous transport inhibition), but highly dependent on K(m1) and V(max1) when dose/EC(50) was (a) <1 (no endogenous transport inhibition) or (b) equal to 1 (with endogenous transport inhibition). Oral input of the endogenous transport inhibitor produced a decrease in t(dur) when the dose/EC(50) range was 0.1-10. These simulations highlight that (a) systemic pharmacokinetic and BBB transport parameters influence t(dur), (b) drug terminal half-life is inversely related to circulating levels of endogenous inhibitors, and (c) oral ingestion of endogenous transport inhibitor(s) reduces t(dur). Overall, these simulations provide insight for the feasibility of rational CNS drug design/delivery via endogenous transporters.