Effects of prenatal exposure to antibodies against brain-specific anion transporter on cognitive functions in rats.

We studied the effects of single intravenous injection of antibodies to brain-specific transmembrane anion transporter (BSAT1; 5 mg/kg) to pregnant rats (gestation day 10) on cognitive functions and behavior of their progeny. One of major functions of BSAT1 (or Oatp1c1) is specific transport of thyroxin across the blood-brain barrier. Female rats of two control groups were injected with non-specific Ig and 0.9% NaCl. The progeny of rats receiving monoclonal antibodies to BSAT1 demonstrated memory impairment in the Y-maze, novel object recognition test, passive avoidance test, and Morris water maze test in comparison with the control group. Our findings suggest that single injection of monoclonal antibodies to BSAT1 during the prenatal period was followed by cognitive impairments, which were probably related to thyroxin deficiency in the nervous tissue.

Organ, cellular, and subcellular localization of brain-specific anion transporter BSAT1.

Organ, cellular, and subcellular localization of brain-specific anion transporter BSAT1 was studied in rats using antibodies to the extracellular fragment (451-557 a.a). The antibodies were shown to recognize the antigen predominantly localized in the nervous tissue, tumors of glial origin, and primordial ovarian follicles. The absence of BSAT1 immunofluorescence signal in kidney and liver sections and accumulation of (125)I labeled antibodies to BSAT1 in these organs indicate that these antibodies do not cross-react with the most common isoforms of OATP expressed in these organs. Analysis of the cellular localization suggests that in the brain, BSAT1 is localized predominantly in astrocytes, but not in endothelial cells, as was previously reported. Laser scanning confocal microscopy with a set of relevant trackers revealed membrane localization of BSAT1. Taking into account the data on the of localization, we can conclude that antibodies to BSAT1 451-557 can be used for basic research of the transport of thyroxin and prostaglandins across the blood brain barrier and for testing the systems for targeted transport of diagnostic preparations and drugs across the blood brain barrier, e.g. to astroglial tumors.

Cloning and expression of brain-specific anion transporter BSAT1 and isolation of monoclonal antibodies to its extracellular fragment.

Brain-specific anion transporter BSAT1 extracellular fragment (451-557) cDNA was cloned in a vector for prokaryotic expression, a producer E. coli strain was obtained, and recombinant extracellular fragment BSAT1(451-557)was purified and used for immunization of BALB/c mice. Splenic cells from mice with verified immune response were used for hybridoma generation. Several hybridoma clones producing monoclonal antibodies to BSAT1 extracellular fragment were selected. Antibody specificity was confirmed by ELISA, immunoblotting with recombinant BSAT1(451-557), and immunofluorescent BSAT1 assay on rat brain sections and cultured HEK293 cells. It was demonstrated that the obtained antibodies specifically bind native rat and human BSAT1 and can be used in both fundamental studies of structures forming the blood-brain barrier and development of targeted transport of diagnostic preparations and drugs across the blood-brain barrier.