Expression of the nuclear transport protein importin β-1 and its association with the neurokinin 3 receptor in the rat hypothalamus following acute hyperosmotic challenge

The tachykinin NK3 receptor (NK3R) is a G protein coupled receptor that is activated, internalized, and trafficked to the nuclei of magnocellular neurons in the paraventricular nucleus of the hypothalamus (PVN) in response to acute hyperosmolarity. The lack of information on the nuclear import pathway raises concerns about the physiological role of nuclear NK3R. NK3R contains a nuclear localizing sequence (NLS) and this raises the possibility that importins are involved in transport of NK3R through the nuclear pore complex. The following experiments utilized: 1) co-immunoprecipitation to determine if NK3R is associated with importin β-1 following activation in response to acute hyperosmolarity in-vivo , and 2) immuno-neutralization of importin β-1 in-vitro to determine if nuclear transport of NK3R was blocked. Rats were given an intravenous injection of hypertonic saline (2 M) and 10 min after the infusion, the PVN was removed and homogenized. Importin β-1 co-immunoprecipitated with the NK3R following treatment with 2 M NaCl, but not following isotonic saline treatment. Immuno-neutralization of importin β-1 decreased the transport of NK3R into the nuclei in a time dependent fashion. The results indicate that in response to acute hyperosmotic challenge, NK3R associates with importin β-1 which enables the nuclear transport of NK3R. This is the first in-vivo study linking importin β-1 and the nuclear transport of a G protein coupled receptor, the NK3R, in brain.