Filchakova O , McIntosh JM .

GM103801 NIGMS NIH HHS , GM48677 NIGMS NIH HHS , P01 GM048677 NIGMS NIH HHS , R01 GM103801 NIGMS NIH HHS

	Figure 2. Comparison between the level of expression of human α9/rat α10 (hα9rα10) and rat α9/human α10 (rα9hα10) receptors.Receptors assembled from injection of cRNAs encoding subunits from different species have different levels of functional expression. hα9rα10 nAChRs were expressed with low efficiency compared to rα9hα10. Results from three batches of oocytes are shown. All oocytes of a given batch were injected on the same day and recordings performed 2 days later. Values of mean current amplitudes are given in Table 1. **p<0.01. Error bars indicate SEM.
	Figure 3. Comparison of functional receptor expression following injection of different ratios of receptor subunit cRNA.Differing subunit ratios of cRNA were injected into oocytes and the resulting levels of expression of functional receptors were compared. Recordings were performed 2 days after injection. The data from oocytes of four different batches were combined to determine the mean current amplitudes. Values are given in Table 2. A one-way ANOVA test with Tukey's post-hoc comparison indicated a significant difference between hα9(1):hα10(1) vs. hα9(5):hα10(1), p<0.001, and between hα9(5):hα10(1) vs. hα9(1):hα10(5), p<0.001. There was no significant difference between hα9(1):hα10(1) and hα9(1):hα10(5), p>0.05.
	Figure 4. Comparison of the 5′ untranslated regions in human α9, human α10, rat α9, and rat α10 subunits.(A) Native 5′UTRs of subunits are between the restriction site and the start codon. (B) The modifications made to the 5′ untranslated region of human α9 and α10 subunits are shown. The 5′UTR of RNA4 of the alfalfa mosaic virus coat protein was inserted into the multiple cloning site of the pSGEM vector between SacII and EcoRI sites. The subunit-encoding sequence is between the EcoRI and XhoI sites.
	Figure 5. AMV improves the level of functional expression of α9-containing nAChRs.(A) Representative traces and (B) comparisons of the levels of functional expression of homomeric human α9 receptors encoded by cRNA without (A, top) and with (A, bottom) AMV. The results are from three different batches of oocytes, each isolated from a different frog and recorded on 3rd day after injection, are presented. (C and D) Comparison of the level of expression of heteromeric receptors. Recordings were conducted on the second day after injection. Values for mean current amplitude are shown in Tables 3 and 4. *p<0.05; **p<0.01. Error bars indicate SEM.
	Figure 1. Comparison between the levels of exogenous expression of rat and human α9-containing nAChRs in X. laevis oocytes.ACh-gated currents were measured in voltage-clamped oocytes as described in Methods. (A) Representative traces from an oocyte injected with human α9 and human α10 cRNA (top) and rat α9 and rat α10 cRNA (bottom). Robust currents were observed with rat cRNA; but only small currents were observed with human cRNA. (B) Comparison of the averaged current responses evoked by 100 µM ACh applications from oocytes expressing human α9α10 and rat α9α10 receptors. The mean current amplitude was 30±3 nA (n = 7 oocytes) for human α9α10 and 8067±1638 nA (n = 7) for rat α9α10, p<0.005. Error bars indicate SEM.

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