Prefrontal beta2 subunit-containing and alpha7 nicotinic acetylcholine receptors differentially control glutamatergic and cholinergic signaling

Abstract

One-second-long increases in prefrontal cholinergic activity ("transients") were demonstrated previously to be necessary for the incorporation of cues into ongoing cognitive processes ("cue detection"). Nicotine and, more robustly, selective agonists at alpha4beta2* nicotinic acetylcholine receptors (nAChRs) enhance cue detection and attentional performance by augmenting prefrontal cholinergic activity. The present experiments determined the role of beta2-containing and alpha7 nAChRs in the generation of prefrontal cholinergic and glutamatergic transients in vivo. Transients were evoked by nicotine, the alpha4beta2* nAChR agonist ABT-089 [2-methyl-3-(2-(S)-pyrrolindinylmethoxy) pyridine dihydrochloride], or the alpha7 nAChR agonist A-582941 [2-methyl-5-(6-phenyl-pyridazin-3-yl)-octahydro-pyrrolo[3,4-c]pyrrole]. Transients were recorded in mice lacking beta2 or alpha7 nAChRs and in rats after removal of thalamic glutamatergic or midbrain dopaminergic inputs to prefrontal cortex. The main results indicate that stimulation of alpha4beta2* nAChRs evokes glutamate release and that the presence of thalamic afferents is necessary for the generation of cholinergic transients. ABT-089-evoked transients were completely abolished in mice lacking beta2* nAChRs. The amplitude, but not the decay rate, of nicotine-evoked transients was reduced by beta2* knock-out. Conversely, in mice lacking the alpha7 nAChR, the decay rate, but not the amplitude, of nicotine-evoked cholinergic and glutamatergic transients was attenuated. Substantiating the role of alpha7 nAChR in controlling the duration of release events, stimulation of alpha7 nAChR produced cholinergic transients that lasted 10- to 15-fold longer than those evoked by nicotine. alpha7 nAChR-evoked cholinergic transients are mediated in part by dopaminergic activity. Prefrontal alpha4beta2* nAChRs play a key role in evoking and facilitating the transient glutamatergic-cholinergic interactions that are necessary for cue detection and attentional performance.

Figure 1.

Choline transients evoked by nicotine and ABT-089 in the prefrontal cortex of C57BL/6J mice. A, Examples of choline transients (self-referenced traces) evoked by pressure ejections of nicotine (1 nmol) and in the presence of TTX (50 pmol; n = 4 per condition). TTX almost completely attenuated the amplitudes of choline transients that were evoked by nicotine (B; mean ± SEM; **p < 0.01). B, Examples of self-referenced traces evoked by pressure ejections of 50 pmol of nicotine or ABT-089. The traces illustrate the greater potency, in terms of amplitude, and the faster decay rate of transients evoked by ABT-089 when compared with nicotine. C, Dose–response curve for the effects of nicotine and ABT-089 on transient amplitudes. Compared with nicotine, ABT-089 was more potent but not more efficacious in increasing cholinergic activity. D, Dose–response curve for the effects of nicotine and ABT-089 on the decay of choline transients. Compared with nicotine, choline transients evoked by ABT-089 returned faster to basal current levels. Nicotine-evoked cholinergic activity lasted longer and more slowly returned to baseline when compared with the more rapid attenuation of ACh release evoked by the α4β2* nAChR agonist (*p < 0.05, **p < 0.01, ***p < 0.001, based on multiple comparisons performed using the LSD for within-subject data and t tests for between-subject data; for significant ANOVA results justifying these multiple comparisons, see Results).

Figure 2.

Choline transients evoked by nicotine (1 nmol) and ABT-089 (1 nmol) in the prefrontal cortex of wild-type and β2 KO mice (the 1 nmol dose for either compound was selected for the assessment of genotype effects because these doses evoke transients with identical amplitudes in the background strain; see Fig. 1D). A, C, Self-referenced traces exemplifying the attenuation of nicotine-evoked (A) or ABT-089-evoked (C) choline transients in β2 KO mice (note that the trace evoked by ABT-089 in β2 KO mice; in C) did not differ from that evoked by vehicle in wild-type mice). B, β2 KO partly attenuated the amplitudes of nicotine-evoked transients and almost completely abolished ABT-089-evoked cholinergic activity. D, β2 KO did not affect the decay rate of nicotine-evoked transients (ABT-089-evoked transients recorded in β2-KO mice were too small to permit reliable determination of t₅₀ values; *p < 0.05, **p <0.01, based on multiple comparisons performed on the basis of significant results of ANOVA; see Results).

Figure 3.

Choline transients evoked by nicotine (1 nmol) and ABT-089 (1 nmol) in the mPFC of wild-type and α7-KO mice. A, B, Representative self-referenced traces depicting choline transients evoked by nicotine (A) or ABT-089 (B) in wild-type and α7 KO mice. The duration but not the amplitudes of nicotine-evoked choline transients was attenuated in α7 KO mice. C, D, ABT-089-evoked transients were not affected by the absence of α7 nAChR (C, D; *p < 0.05, based on multiple comparisons performed on the basis of significant results of ANOVA; see Results).

Figure 4.

nAChR agonist-evoked glutamatergic transients in the mPFC of wild-type and β2 KO mice. A, B, In contrast to glutamate released measured by using conventional in vivo microdialysis methods, blockade of voltage-regulated sodium channels by TTX almost completely abolished nicotine-evoked increases in glutamate release. C, With respect to the amplitudes of glutamatergic transients, ABT-089 was more potent but not more efficacious than nicotine. D, The decay rate of glutamatergic transients evoked by ABT-089 remained flat over the entire dose range; in contrast, increasing doses of nicotine produced transients that required increasingly more time to decay, yielding a significantly slower decay of nicotine-evoked glutamatergic transients in response to the highest dose of drug. E, In β2 KO mice, the amplitudes of glutamatergic transients evoked by nicotine (1 nmol) were partly attenuated, whereas ABT-089 (1 nmol)-evoked transients were completely abolished. F, The decay of nicotine-evoked transients appeared to be slower in β2 KO mice, but this difference did not reach significance (ABT-089-evoked glutamatergic transients remained too small to allow a reliable determination of t₅₀ values; *p < 0.05, **p < 0.01, ***p < 0.001, respectively, based on multiple comparisons performed using the LSD for within-subject data and t tests for between-subject data; for significant ANOVA results justifying these multiple comparisons, see Results).

Figure 5.

nAChR agonist-evoked glutamatergic transients in the mPFC of wild-type and α7 KO mice. A, B, Examples of glutamatergic transients evoked by nicotine (A) or ABT-089 (B) in wild-type and α7 KO mice. Knock-out of α7 nAChRs reduced the duration, but did not affect the amplitude, of transients evoked by nicotine. C, D, ABT-089-evoked transients were not affected by α7 KO (C, D; *p < 0.05, based on multiple comparisons performed on the basis of significant results of ANOVA; see Results).

Figure 6.

Effects of lesions of the MD on prefrontal choline transients evoked by the α4β2* nAChR agonist ABT-089. A shows a Nissl-stained coronal section from an intact brain depicting the area of the MD as well as the adjacent nuclei [central medial thalamic nucleus (CM), oral paracentral thalamic nucleus (OPC), posterior paraventricular nucleus (PVP), and habenular]. B, Infusions of ibotenic acid produced extensive gliosis in the area of the MD; arrows depict the borders of the region lacking neurons. C, Self-referenced choline transients evoked by the α4β2* nAChR agonist in control and lesioned animals, respectively.

Figure 7.

Cholinergic transients evoked by prefrontal pressure ejections of the α7 nAChR agonist A-582941 and role of dopaminergic afferents. A, Examples of traces evoked by 200 pmol and 2 nmol of A-582941. Note the extremely long-lasting increases in ACh release evoked by the α7 nAChR agonist; these release events began to decline 100–150 s after the administration of the higher dose, contrasting with 3–5 s for transients evoked by α4β2* nAChR agonists and with tens of seconds for transients evoked by nicotine (see Fig. 1; for t₅₀ values for transients evoked by A-582941, see Results). Coadministration of the α7 nAChR antagonist MLA attenuated transients evoked by A-582941 (see B for effects on peak amplitudes). C, Removal of dopaminergic inputs to the PFC partly attenuated the release evoked by the α7 nAChR agonist. C shows examples of cholinergic transients evoked by α7 nAChR agonist A-582941 (2 nmol) in a sham-operated control animal and after dopaminergic lesions (for histological documentation of these lesions see Fig. 8; *p < 0.05).

Figure 8.

Lesions of the midbrain dopaminergic system. A–D are coronal sections immunostained against TH. A, Prelimbic region from the brain of an intact rat. TH-immunoreactive fibers and varicosities are visible throughout all layers, with vertical fibers being prominent in deep layers and white matter. C, In lesioned animals, TH-immunopositive puncta are almost completely abolished (see Results for quantification); residual TH-positive fibers were observed in deep layers and white matter (arrows). B, Substantia nigra (SN) and VTA dopaminergic neurons in an intact brain. D, The lesions almost completely destroyed TH-positive neurons in the midbrain (see arrows for residual neurons).