Imaging cationic drug H+/antiporter function at the blood-brain barrier using 11C-diphenhydramine PET imaging

Objectives Evidence for a cationic drug H+/antiporter of the solute carrier transporter (SLC) family, also known as clonidine antiporter, has been functionally demonstrated at the blood-brain barrier (BBB) using invasive approaches. This transporter imports many psychoactive drugs into the brain, including clonidine, oxycodone, nicotine, cocaine and diphenhydramine, a sedative H1-histamine receptor antagonist. Drug H+/antiporter function at the BBB challenges the assumption that only passive diffusion controls the distribution of drugs to the central nervous system (CNS). Non-invasive tools are required to identify safe and potent in vivo inhibitors and study the pathophysiological regulation of H+/antiporter function in animals and humans. In the present study, we developed and validated 11C-diphenhydramine as a PET radioligand to estimate H+/antiporter function at the rat BBB.

Methods Diphenhydramine was isotopically radiolabeled using carbon-11 and standard radiomethylation of its N-desmethyl precursor. 11C-diphenhydramine microPET imaging was performed in 4 rats for 60 min. Brain and plasma radiometabolites were studied using radio-HPLC to measure a metabolite-corrected input function. The initial brain uptake clearance (Cluptake) of 11C-diphenhydramine was estimated using the integration plot method from 0 to 2 min. Displacement experiments using 1 mg/kg diphenhydramine injected 30 min after 11C-diphenhydramine were performed in 3 other rats to investigate the contribution of 11C-diphenhydramine specific binding to the brain. Because higher dose diphenhydramine is lethal, saturation experiments were performed using in situ brain perfusion in rats to i) assess the respective contribution of passive and carrier-mediated diffusion to the BBB permeation of diphenhydramine and ii) assess the concentration dependency of the 14C-diphenhydramine uptake clearance (Kin) in the living brain.

Results 11C-diphenhydramine brain uptake was substantial and homogeneously distributed within the brain parenchyma. Displacement using 1 mg/kg diphenhydramine had no impact on radioactivity brain kinetics. 11C-diphenhydramine radiometabolites accounted for 72 % and 76 % in plasma and for <2 % and 20 % in the brain at 15 min and 60 min, respectively. PET-based estimation of brain 11C-diphenhydramine Cluptake was 0.99 ± 0.18 mL.min-1.cm-3. 14C-diphenhydramine in situ brain perfusion led to a baseline Kin of 1.8 ± 0.1 mL.min-1g-1 and unveiled a saturable transport of high capacity (Km = 4 mM) across the BBB. The carrier-mediated component accounted for ~75% of diphenhydramine total influx through the BBB in its therapeutic concentration range (1-2 µM).

Conclusions Diphenhydramine uptake by the brain is mainly controlled by saturable influx through the BBB, suggesting major impact of H+/antiporter activity at the BBB compared to passive diffusion. 11C-diphenhydramine binding to the brain is non-specific (non-displaceable). In the absence of brain radiometabolites in the first 15 min scanning, 11C-diphenhydramine brain uptake clearance can be reliably estimated using PET. Cluptake value is in the same order of magnitude as Kin calculated using in situ brain perfusion. 11C-diphenhydramine benefits from favorable kinetic properties and offers a suitable imaging tool for the non-invasive study of this H+/antiporter function at the BBB using PET. $$graphic_7F7D7378-5097-4821-9428-0AAA08A65CC2$$