Nicotine addiction relies on brain receptors that have been difficult to fully study and characterize. Scientists at the University of Colorado in Boulder have demonstrated that an immunolabeling technique can effectively analyze receptor subunits.

Background: Nicotine’s effects on the brain are triggered upon its binding to nicotinic acetylcholine receptors, each of which consists of five subunits: two alphas, one beta, one delta and one gamma. Different combinations of these subunits produce different receptor subtypes, which may vary in their pharmacology, biophysical properties, and distribution. To more fully understand how to interfere with nicotine’s effects in the brain, scientists must first understand where these different receptors are and how they work. Two of the most important subunits, a4 and b2, have been hard to study because current study methods can only locate the fully assembled receptor unit. Researchers wanted to know if an alternative strategy of immunolabeling (i.e., using antibodies to tag individual proteins), which has been fraught with technical challenges, would be able to identify, map, and quantify separate subunits.

Study Design: Scientists at the University of Colorado worked with brain sections of mice genetically engineered to express particular a4 and b2 subunit combinations. Using a sensitive immunolabeling technique, they explored the expression of the a4 and b2 subunits at both the gene and protein levels. Additional mice strains, missing the subunits under study, were used as controls.

What They Found: The two predominant nicotinic receptor subtypes (a4 and b2) were reliably detected using immunolabeling. Expression of the a4 subunit protein was almost universally dependent on b2, whereas most, but not all, b2 subunit protein expression was a4-dependent.

Comments from the Authors: Immunolabeling using specific antibodies offers a powerful approach for mapping the distribution of nicotine receptor subunits and can produce reliable quantitative results.

What’s Next: Similar studies can be designed to locate other nicotine receptor subtypes. In many cases, the antibody recognition sites are inside the cell membrane. It will likely take alternative biochemical approaches to uncover these less accessible sites. A better understanding of receptor composition and function may eventually have important implications for developing interventions at the receptor level.

Source: The study, led by Dr. Paul Whiteaker of the Institute for Behavioral Genetics at the University of Colorado, Boulder, with Dr. Jon Lindstrom of the University of Pennsylvania, was published in volume 499, number 6, pages 1016-1038 (2006) of the Journal of Comparative Neurology.