Inhibition of the Jak-STAT pathway prevents CNTF-mediated survival of axotomized oxytocinergic magnocellular neurons in organotypic cultures of the rat supraoptic nucleus

Abstract

Previous studies have demonstrated that ciliary neurotrophic factor (CNTF) enhances survival and process outgrowth from magnocellular neurons in the paraventricular (PVN) and the supraoptic (SON) nuclei. However, the mechanisms by which CNTF facilitates these processes remain to be determined. Therefore, the aim of this study was to identify the immediate signal transduction events that occur within the rat SON following administration of exogenous rat recombinant CNTF (rrCNTF) and to determine the contribution of those intracellular signaling pathway(s) to neuronal survival and process outgrowth, respectively. Immunohistochemical and Western blot analysis demonstrated that axonal injury and acute unilateral pressure injection of 100 ng/μl of rrCNTF directly over the rat SON resulted in a rapid and transient increase in phosphorylated-STAT3 (pSTAT3) in astrocytes but not neurons in the SON in vivo. Utilizing rat hypothalamic organotypic explant cultures, we then demonstrated that administration of 25 ng/ml rrCNTF for 14 days significantly increased the survival and process outgrowth of OT magnocellular neurons. In addition, pharmacological inhibition of the Jak-STAT pathway via AG490 and cucurbitacin I significantly reduced the survival of OT magnocellular neurons in the SON and PVN; however, the contribution of the Jak-STAT pathway to CNTF-mediated process outgrowth remains to be determined. Together, these data indicate that CNTF-induced survival of OT magnocellular neurons is mediated indirectly through astrocytes via the Jak-STAT signaling pathway.

Inhibition of a novel specific neuroglial integrin signaling pathway increases STAT3-mediated CNTF expression

“…Conversely, the CNTF-stimulatory pathway is activated by neural cytokines such as CNTF, LIF and IL-6 which bind to their respective gp130 (gp) receptor complexes (CNTF: CNTFαR/LIFβR/gp130; LIF: LIFβR/gp130; IL-6: IL-6αR/gp130/gp130; Lβ indicates LIFβR), triggering recruitment of JAK to activate STAT3 by phosphorylation at the pro-transcription residue tyrosine-705, y705; Green). CNTF is known to activate STAT3 in astrocytes in vitro [9,87-90] and in vivo [91]. FAK may simultaneously stimulate the activating pathway, potentially representing another component of the tightly regulated CNTF gene expression under physiological conditions.…”

“…It is possible that pSTAT3 (Ser-727) has gene specific interactions similar to methyl CpG binding protein 2 which can inhibit [85] or activate transcription when associated with other transcription factors [86]. In astrocytes, CNTF induces phosphorylation of STAT3 at Tyr-705 for transcriptional activity in vitro [9,87-90] and in vivo [91]. C6 glioma cells reportedly do not express the CNTF alpha receptor [92] but can respond to CNTF [93], possibly through the IL-6 receptor to activate JAK-STAT3 (Tyr-705) signaling as shown in BaF3 cells [94].…”

“…With such diverse functions and as a mediator of critical protective STAT3 signaling in neurons [90], it is likely that several molecular mechanisms exist that lead to CNTF transcription.…”

Absence of axonal sprouting following unilateral lesion in 125‐day‐old rat supraoptic nucleus may be due to age‐dependent decrease in protein levels of ciliary neurotrophic factor receptor alpha

“…We have hypothesized that the collateral sprouting response following unilateral lesion is due to a protein that is inherent to astrocytes in the SON, ciliary neurotrophic factor (CNTF). In hypothalamic organotypic cultures, CNTF promotes neuron survival (Askvig & Watt, ; Askvig et al, ; Rusnak, House, Arima, & Gainer, ; Vutskits, Bartanusz, Schulz, & Kiss, ) and axonal sprouting (Askvig & Watt, ) following axotomy. Additionally, CNTF protein levels increase in the SON during the collateral sprouting response (Askvig, Leiphon, & Watt, ; Watt, Bone, Pressler, Cranston, & Paden, ), further suggesting that it may play a role in promoting the axonal sprouting from uninjured neurons following unilateral lesion.…”

The MAPK and PI3K pathways mediate CNTF-induced neuronal survival and process outgrowth in hypothalamic organotypic cultures

“…However, there is growing evidence indicating that ciliary neurotrophic factor (CNTF) plays an important role. For example, CNTF has been shown to promote axonal sprouting of hypothalamic magnocellular neurons in vitro (Askvig et al 2013 ; Vutskits et al 1998 ) and motor neuron sprouting in vivo (Gurney et al 1992 ; Guthrie et al 1997 ; Kwon and Gurney 1994 ; Oyesiku and Wigston 1996 ; Siegel et al 2000 ; Simon et al 2010 ; Ulenkate et al 1994 ; Wright et al 2007 ; Xu et al 2009 ). Moreover, CNTF promotes the survival of magnocellular neurons in the paraventricular nucleus (PVN) and supraoptic nucleus (SON) of the magnocellular neurosecretory system in organotypic cultures (Askvig et al 2013 ; House et al 2009 ; Rusnak et al 2002 ; Rusnak et al 2003 ; Vutskits et al 1998 ; Vutskits et al 2003 ).…”