Influence of Subunit Composition on Desensitization of Neuronal Acetylcholine Receptors at Low Concentrations of Nicotine

Abstract

The influence of ␣ and ␤ subunits on the properties of nicotineinduced activation and desensitization of neuronal nicotinic acetylcholine receptors (nAChRs) expressed in Xenopus oocytes was examined. Receptors containing ␣4 subunits were more sensitive to activation by nicotine than ␣3-containing receptors. At low concentrations of nicotine, nAChRs containing ␤2 subunits reached near-maximal desensitization more rapidly than ␤4-containing receptors. The concentration of nicotine producing half-maximal desensitization was influenced by the particular ␣ subunit expressed; similar to results for activation, ␣4-containing receptors were more sensitive to desensitizing levels of nicotine than ␣3-containing receptors. The ␣ subunit also influenced the rate of recovery from desensitization; this rate was approximately inversely proportional to the apparent nicotine affinity for the desensitized state. The homomeric ␣7 receptor showed the lowest sensitivity to nicotine for both activation and desensitization; ␣7 nAChRs also demonstrated the fastest desensitization kinetics. These subunitdependent properties remained in the presence of external calcium, although subtle, receptor subtype-specific effects on both the apparent affinities for activation and desensitization and the desensitization kinetics were noted. These data imply that the subunit composition of various nAChRs determines the degree to which receptors are desensitized and/or activated by tobacco-related levels of nicotine. The subtype-specific balance between receptor activation and desensitization should be considered important when the cellular and behavioral actions of nicotine are interpreted.

Desensitization of neuronal nicotinic receptors

“…Moreover, functional studies designed to replicate the radiolabeled ligand binding assays find that ␣4 subunit-containing nAChRs are desensitized by much lower concentrations of agonist than those containing ␣3 subunits. The IC 50 values for desensi-tization of functional channels by nicotine/ACh are in the same range as their binding K d 's, Ϸ0.1-10 nM and Ϸ150 -400 nM for ␣4* and ␣3* nAChRs, respectively (Hsu et al, 1996;Fenster et al, 1997), thus confirming the difference in affinity of the "D" state for these receptors. In addition, chimeric ␣3/␣4 subunits (containing the N-terminal agonist binding domain of ␣3 subunits), when expressed with ␤2 subunits, create receptors with an ␣3-like desensitization affinity (IC 50 Ϸ 700 nM) compared to ␣4␤2 (IC 50 Ϸ 50 nM), implying that the ␣ subunit is the primary determinant of affinity (Kuryatov et al, 2000).…”

“…2). Likewise, it is probable that desensitization of the lower affinity putative ␣3␤4*-nAChRs, as observed in various cells representative of both the autonomic nervous system and CNS (Higgins and Berg, 1988;Oortigiesen and Vijverberg, 1989;Mathie et al, 1990;Khiroug et al, 1997Khiroug et al, , 1998Boyd, 1987;Ifune and Steinbach, 1993;Lester and Dani, 1995), in addition to systems designed to specifically express ␣3␤4 receptors (Cachelin and Jaggi, 1991;Hsu et al, 1996;Fenster et al, 1997;Wang et al, 1998;Xiao et al, 1998), can also be explained in terms of the two-state model, although due to the slower time course of desensitization for ␤4 subunit-containing receptors (see below), the biphasic nature may only be seen clearly during longer agonist applications (e.g., Lester and Dani, 1995). Concentration-dependent analyses of desensitization of presumed native ␣3␤4* nAChRs have estimated the affinity of the desensitized state for nicotine to be Ϸ20 -300 nM (Higgins and Berg, 1988;Lester and Dani, 1995).…”

“…Following on from the work of Cachelin and Jaggi (1991), several studies have suggested that the ␤ subunit (␤4/␤2) strongly influences the rate of onset of desensitization: receptors containing ␤2 subunits desensitize faster than those containing ␤4 subunits (Hsu et al, 1995;Fenster et al, 1997;Wang et al, 1998;Bohler et al, 2001). The difference is effectively due to an altered balance of the fast and slow desensitization components: Ͼ90% desensitization for ␤4*nAChRs is slow; Ͼ90% desensitization for ␤2*nAChRs is fast ( Fig.…”

“…3; Bohler et al, 2001). Arguments for the dominant role of the ␤ subunit in desensitization onset kinetics are strengthened by the findings that this characteristic is independent of the type of ␣ subunit expressed (␣3, ␣4, Fenster et al, 1997; ␣6, Kuryatov et al, 2000), whether nicotine or ACh is used (Fenster et al, 1997;Bohler et al, 2001), and despite which cell type is studied . Conversely, in experiments in which the ␤ subunit is kept constant, receptors containing ␣3 subunits desensitize faster than those containing ␣4 (Gross et al, 1991;Vibat et al, 1995;Chavez-Noriega et al, 2000).…”

“…Conversely, in experiments in which the ␤ subunit is kept constant, receptors containing ␣3 subunits desensitize faster than those containing ␣4 (Gross et al, 1991;Vibat et al, 1995;Chavez-Noriega et al, 2000). Thus, overall heteromeric nAChRs can be ordered in terms of their desensitization kinetics (from fastest to slowest): ␣3␤2 Ͼ ␣4␤2 Ͼ ␣3␤4 Ͼ ␣4␤4 (see Fenster et al, 1997). There has so far been little interest in the desensitization contributions of ␣2 subunits, perhaps due to their restricted CNS distribution (Wada et al, 1989; but see Zoli et al, 1998;Cordero-Erausquin et al, 2000).…”