title: Positive allosteric modulation of alpha-7 nicotinic receptors promotes cell death by inducing Ca(2+) release from the endoplasmic reticulum.
abstract: Positive allosteric modulation of α7 isoform of nicotinic acetylcholine receptors (α7-nAChRs) is emerging as a promising therapeutic approach for central nervous system disorders such as schizophrenia or Alzheimer's disease. However, its effect on Ca(2+) signaling and cell viability remains controversial. This study focuses on how the type II positive allosteric modulator (PAM II) PNU120596 affects intracellular Ca(2+) signaling and cell viability. We used human SH-SY5Y neuroblastoma cells overexpressing α7-nAChRs (α7-SH) and their control (C-SH). We monitored cytoplasmic and endoplasmic reticulum (ER) Ca(2+) with Fura-2 and the genetically encoded cameleon targeting the ER, respectively. Nicotinic inward currents were measured using patch-clamp techniques. Viability was assessed using methylthiazolyl blue tetrazolium bromide or propidium iodide staining. We observed that in the presence of a nicotinic agonist, PNU120596 (i) reduced viability of α7-SH but not of C-SH cells; (ii) significantly increased inward nicotinic currents and cytosolic Ca(2+) concentration; (iii) released Ca(2+) from the ER by a Ca(2+) -induced Ca(2+) release mechanism only in α7-SH cells; (iv) was cytotoxic in rat organotypic hippocampal slice cultures; and, lastly, all these effects were prevented by selective blockade of α7-nAChRs, ryanodine receptors, or IP3 receptors. In conclusion, positive allosteric modulation of α7-nAChRs with the PAM II PNU120596 can lead to dysregulation of ER Ca(2+) , overloading of intracellular Ca(2+) , and neuronal cell death. This study focuses on how the type II positive allosteric modulator PNU120596 (PAM II PNU12) affects intracellular Ca(2+) signaling and cell viability. Using SH-SY5Y neuroblastoma cells overexpressing α7-nAChRs (α7-SH) and their control (C-SH), we find that PAM of α7-nAChRs with PNU120596: (i) increases inward calcium current (ICa ) and cytosolic Ca(2+) concentration ([Ca(2+) ]cyt ); (ii) releases Ca(2+) from the ER ([Ca(2+) ]ER ) by a Ca(2+) -induced Ca(2+) release mechanism; and (iv) reduces cell viability. These findings were corroborated in rat hippocampal organotypic cultures. [Ca(2+) ]cyt , cytosolic Ca(2+) concentration; [Ca(2+) ]ER , endoplasmic reticulum Ca(2+) concentration; α7 nAChR, α7 isoform of nicotinic acetylcholine receptors; α7-SH, SH-SY5Y stably overexpressing α7 nAChRs cells; C-SH, control SH-SY5Y cells; Nic, nicotine; PNU12, PNU120596.
authors: Servicio de Farmacología Clínica, Instituto de Investigación Sanitaria, Hospital Universitario de la Princesa, Madrid, Spain.
authors: María
authors: M
authors: Guerra-Álvarez
authors: Ana J
authors: AJ
authors: Moreno-Ortega
authors: Elisa
authors: E
authors: Navarro
authors: José Carlos
authors: JC
authors: Fernández-Morales
authors: Javier
authors: J
authors: Egea
authors: Manuela G
authors: MG
authors: López
authors: María F
authors: MF
authors: Cano-Abad
mesh_terms: D000494:Allosteric Regulation / Q000187:drug effects / Q000502:physiology; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D016923:Cell Death / Q000187:drug effects / Q000502:physiology; D045744:Cell Line, Tumor; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D007555:Isoxazoles / Q000494:pharmacology; D008297:Male; D010671:Phenylurea Compounds / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D064569:alpha7 Nicotinic Acetylcholine Receptor / Q000819:agonists* / Q000502:physiology*
chemical_list: C508388:1-(5-chloro-2,4-dimethoxyphenyl)-3-(5-methylisoxazol-3-yl)urea; D007555:Isoxazoles; D010671:Phenylurea Compounds; D064569:alpha7 Nicotinic Acetylcholine Receptor; D002118:Calcium
keywords: Ca2+; D1-ER cameleon probe; cytotoxicity; positive allosteric modulators; α7 nicotinic receptors