Promises and pitfalls of a Pannexin1 transgenic mouse line

Abstract

Gene targeting strategies have become a powerful technology for elucidating mammalian gene function. The recently generated knockout (KO)-first strategy produces a KO at the RNA processing level and also allows for the generation of conditional KO alleles by combining FLP/FRT and Cre/loxP systems, thereby providing high flexibility in gene manipulation. However, this multipurpose KO-first cassette might produce hypomorphic rather than complete KOs if the RNA processing module is bypassed. Moreover, the generation of a conditional phenotype is also dependent on specific activity of Cre recombinase. Here, we report the use of an efficient molecular biological approach to test pannexin1 (Panx1) mRNA expression in global and conditional Panx1 KO mice derived from the KO-first mouse line, Panx1tm1a(KOMP)Wtsi. Using qRT-PCR, we demonstrate that tissues from wild-type (WT) mice show a range of Panx1 mRNA expression levels, with highest expression in trigeminal ganglia, bladder and spleen. Unexpectedly, we found that in mice homozygous for the KO-first allele, Panx1 mRNA expression is not abolished but reduced by 70% compared to that of WT tissues. Thus, Panx1 KO-first mice present a hypomorphic phenotype. Crosses of Panx1 KO-first with FLP deleter mice generated Panx1f/f mice. Further crosses of the latter mice with mGFAP-Cre or NFH-Cre mice were used to generate astrocyte- and neuron-specific Panx1 deletions, respectively. A high incidence of ectopic Cre expression was found in offspring of both types of conditional Panx1 KO mice. Our study demonstrates that Panx1 expression levels in the global and conditional Panx1 KO mice derived from KO-first mouse lines must be carefully characterized to ensure modulation of Panx1 gene expression. The precise quantitation of Panx1 expression and its relation to function is expected to provide a foundation for future efforts aimed at deciphering the role of Panx1 under physiological and pathological conditions.

Glial pannexin1 contributes to tactile hypersensitivity in a mouse model of orofacial pain

“…Panx1 channels are present in both neurons and satellite glial cells in trigeminal ganglia of mice10 (Suppl. Fig.…”

“…In the nervous system, Panx1 channels are expressed in neurons and glia78910; upon activation, Panx1 channels release the algogenic molecule ATP and functionally and structurally associate with components of the inflammasome leading to cytokine production91112. Evidence from animal models indicates that Panx1 plays a pivotal role in several types of pain, including trigeminal hypersensitivity413, spared nerve injury- and chemotherapy-induced neuropathic pain141516, and migraine17.…”

Restoration of Connexin 40 (Cx40) in Renin-Producing Cells Reduces the Hypertension of Cx40 Null Mice

“…17 These differences may be because of a partial, and possibly not fully cell-specific loss of Cx40, which in this study was solely supported by negative immunofluorescence observations, 17,26,27 as sometimes observed with the Cre-lox methods. 28,29 Also, this study 17 did not investigate the effects of the Cx40 deletion on other proteins of RSCs and ECs, 12,13,17,18 thus preventing the unambiguous identification of the biological effects specifically attributable to Cx40. To address these issues, we have generated 2 novel mouse lines in which Cx40 was selectively restored in either the ECs or the RSCs of Cx40 −/− mice.…”

Exciting and not so exciting roles of pannexins

“…The distribution of Panx1 is widespread, being present in most types of cells and tissues [2, 11, 25]. In the nervous system, Panx1 is prominently expressed in the CNS (cerebellum, hippocampus, olfactory bulb, and thalamus: [26–28]) and in the PNS (sensory ganglia: [25, 29, 30]). In the hippocampus, high Panx1 expression was found in GABAergic and glutamatergic neurons within the dentate gyrus and all CA regions, stratum oriens, stratum radiatum, and stratum lacunosum-moleculare [28].…”