Rat neuronal nicotinic acetylcholine receptors containing α7 subunit: pharmacological properties of ligand binding and function
Abstract
Aim: To compare pharmacological properties of heterologously expressed homomeric α7 nicotinic acetylcholine receptors (α7 nAChRs) with those of native nAChRs containing α7 subunit (α7* nAChRs) in rat hippocampus and cerebral cortex.
Methods: We established a stably transfected HEK-293 cell line that expresses homomeric rat α7 nAChRs. We studies ligand binding profiles and functional properties of nAChRs expressed in this cell line and native rat α7*nAChRs in rat hippocampus and cerebral cortex. We used [125I]-α-bungarotoxin to compare ligand binding profiles in these cells with those in rat hippocampus and cerebral cortex. The functional properties of the α7 nAChRs expressed in this cell line were studied using whole-cell current recording.
Results: The newly established cell line, KXα7R1, expresses homomeric α7 nAChRs that bind [125I]-α-bungarotoxin with a Kd value of 0.38±0.06 nmol/L, similar to Kd values of native rat α7* nAChRs from hippocampus (Kd=0.28±0.03 nmol/L) and cerebral cortex (Kd=0.33±0.05 nmol/L). Using whole-cell current recording, the homomeric α7 nAChRs expressed in the cells were activated by acetylcholine and (−)-nicotine with EC50 values of 280±19 μmol/L and 180±40 μmol/L, respectively. The acetylcholine activated currents were potently blocked by two selective antagonists ofα7 nAChRs, α-bungarotoxin (IC50=19±2 nmol/L) and methyllycaconitine (IC50=100±10 pmol/L). A comparative study of ligand binding profiles, using 13 nicotinic ligands, showed many similarities between the homomeric α7 nAChRs and native α7* receptors in rat brain, but it also revealed several notable differences.
Conclusion: This newly established stable cell line should be very useful for studying the properties of homomeric α7 nAChRs and comparing these properties to native α7* nAChRs.
Keywords:
Methods: We established a stably transfected HEK-293 cell line that expresses homomeric rat α7 nAChRs. We studies ligand binding profiles and functional properties of nAChRs expressed in this cell line and native rat α7*nAChRs in rat hippocampus and cerebral cortex. We used [125I]-α-bungarotoxin to compare ligand binding profiles in these cells with those in rat hippocampus and cerebral cortex. The functional properties of the α7 nAChRs expressed in this cell line were studied using whole-cell current recording.
Results: The newly established cell line, KXα7R1, expresses homomeric α7 nAChRs that bind [125I]-α-bungarotoxin with a Kd value of 0.38±0.06 nmol/L, similar to Kd values of native rat α7* nAChRs from hippocampus (Kd=0.28±0.03 nmol/L) and cerebral cortex (Kd=0.33±0.05 nmol/L). Using whole-cell current recording, the homomeric α7 nAChRs expressed in the cells were activated by acetylcholine and (−)-nicotine with EC50 values of 280±19 μmol/L and 180±40 μmol/L, respectively. The acetylcholine activated currents were potently blocked by two selective antagonists ofα7 nAChRs, α-bungarotoxin (IC50=19±2 nmol/L) and methyllycaconitine (IC50=100±10 pmol/L). A comparative study of ligand binding profiles, using 13 nicotinic ligands, showed many similarities between the homomeric α7 nAChRs and native α7* receptors in rat brain, but it also revealed several notable differences.
Conclusion: This newly established stable cell line should be very useful for studying the properties of homomeric α7 nAChRs and comparing these properties to native α7* nAChRs.