Characterization of a novel brain barrier ex vivo insect‐based P‐glycoprotein screening model

Abstract

In earlier studies insects were proposed as suitable models for vertebrate blood–brain barrier (BBB) permeability prediction and useful in early drug discovery. Here we provide transcriptome and functional data demonstrating the presence of a P-glycoprotein (Pgp) efflux transporter in the brain barrier of the desert locust (Schistocerca gregaria). In an in vivo study on the locust, we found an increased uptake of the two well-known Pgp substrates, rhodamine 123 and loperamide after co-administration with the Pgp inhibitors cyclosporine A or verapamil. Furthermore, ex vivo studies on isolated locust brains demonstrated differences in permeation of high and low permeability compounds. The vertebrate Pgp inhibitor verapamil did not affect the uptake of passively diffusing compounds but significantly increased the brain uptake of Pgp substrates in the ex vivo model. In addition, studies at 2°C and 30°C showed differences in brain uptake between Pgp-effluxed and passively diffusing compounds. The transcriptome data show a high degree of sequence identity of the locust Pgp transporter protein sequences to the human Pgp sequence (37%), as well as the presence of conserved domains. As in vertebrates, the locust brain–barrier function is morphologically confined to one specific cell layer and by using a whole-brain ex vivo drug exposure technique our locust model may retain the major cues that maintain and modulate the physiological function of the brain barrier. We show that the locust model has the potential to act as a robust and convenient model for assessing BBB permeability in early drug discovery.

Transepithelial transport of P-glycoprotein substrate by the Malpighian tubules of the desert locust

“…Secondly, the addition of verapamil, a P-glycoprotein inhibitor (e.g. [23,27,28,39]), significantly reduced the extrusion of rhodamine B. The presence of P-glycoproteins in other tissues of the desert locusts also supports our conclusion that they are expressed in the Malpighian tubules; proteins with a comparable physiology and similar sequence to the human P-glycoprotein (ABCB1 gene) are expressed within the blood brain barrier of the locusts S .…”

“…migratoria , P . americana ) from orthopteroid orders [15]; and (2) they are expressed in the blood brain barrier of the desert locust [23]. However, P-glycoproteins in the Malpighian tubules of desert locusts have not been studied previously.…”

“…[25,26]) with or without the addition of the selective P-glycoprotein inhibitor verapamil (e.g. [23,27,28]). Our results suggest that P-glycoprotein transporters may play an important role in the xenobiotic extrusion in the Malpighian tubules of the desert locust.…”

Multi-Drug Resistance Transporters and a Mechanism-Based Strategy for Assessing Risks of Pesticide Combinations to Honey Bees

“…Therefore, we expect hemolymph concentrations of RhB to be higher in bees treated with transporter inhibitors as a result of reduced transporter function in the midgut and Malpighian tubules [ 45 , 53 – 56 ]. We also expect higher RhB concentrations in the heads (composed largely of brain and salivary glands) of bees treated with inhibitors because, in addition to higher hemolymph-mediated exposure, these transporters have key roles in maintaining the blood-brain barrier in insects, preventing the accumulation of xenobiotics in brain tissue [ 43 , 44 , 50 ].…”

Desenvolvimento de métodos de geração de imagem por espectrometria de massas (MALDI e DESI-MSI) aplicados a modelos in vitro e in vivo de permeação cutânea e sistema nervoso central

“…Com isso, podemos concluir que a técnica DESI-MSI aplicada a estudos de metabolismo em modelo de inseto pode ser uma ferramenta interessante (OLSEN et al, 2014;OLSEN et al, 2015). No entanto DESI-MSI trata-se de uma técnica ainda pouco explorada, assim como o modelo de inseto, ainda muito recente e com poucos relatos na literatura.…”