July 2018
Volume 59, Issue 9
ARVO Annual Meeting Abstract  |   July 2018
Nuclear Cyclic-AMP Signaling in Developing Astrocytes
Author Affiliations & Notes
  • Evan G. Cameron
    Stanford University, Palo Alto, California, United States
  • Sahil Shah
    Stanford University, Palo Alto, California, United States
  • Joana Galvao
    Stanford University, Palo Alto, California, United States
  • Michael Nahmou
    Stanford University, Palo Alto, California, United States
  • Masoumeh Ashouri
    Stanford University, Palo Alto, California, United States
  • Michael Kapiloff
    Stanford University, Palo Alto, California, United States
  • Jeffrey L Goldberg
    Stanford University, Palo Alto, California, United States
  • Footnotes
    Commercial Relationships   Evan Cameron, None; Sahil Shah, None; Joana Galvao, None; Michael Nahmou, None; Masoumeh Ashouri, None; Michael Kapiloff, None; Jeffrey Goldberg, None
  • Footnotes
    Support  F32-EY025915-04, P30-EY026877
Investigative Ophthalmology & Visual Science July 2018, Vol.59, 1484. doi:https://doi.org/
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      Evan G. Cameron, Sahil Shah, Joana Galvao, Michael Nahmou, Masoumeh Ashouri, Michael Kapiloff, Jeffrey L Goldberg; Nuclear Cyclic-AMP Signaling in Developing Astrocytes. Invest. Ophthalmol. Vis. Sci. 2018;59(9):1484. doi: https://doi.org/.

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      © ARVO (1962-2015); The Authors (2016-present)

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Purpose : How does compartmentalized cyclic-AMP (cAMP) signaling influence glial cell development in the central nervous system (CNS)? Astrocytes are the principal glial cell in the mammalian brain that provide trophic and metabolic support to neurons, maintain ion homeostasis, buffer excess neurotransmitters and respond to injury. As such, proper astrocyte development is critical to CNS function. Many processes associated with astrocyte development are regulated by cAMP including proliferation, differentiation and migration. The source, extent and subcellular domains relevant to these cAMP-mediated signals, however, remain poorly understood. Here, we report that nuclear localized cAMP derived from a calcium-sensitive soluble adenylyl cyclase (sAC) is necessary for proliferation in developing astrocytes.

Methods : Primary cortical astrocytes were purified from E16 mouse pups for in vitro studies. Astrocyte proliferation was assessed by EdU incorporation and cell cycle progression measured using propidium iodide. Plasmids encoding cAMP-sponges were transfected using lipofectamine. KH7, 2HE and LRE1 were used to pharmacologically inhibit sAC activity. sAC knockout experiments were carried out in primary sACfloxed astrocytes treated with lentiviral-Cre.

Results : To determine if nuclear localized cAMP plays a role in astrocyte proliferation, we targeted PKA-derived cAMP “sponges” to various subcellular compartments to inhibit local cAMP signaling. Interestingly, we found buffering nuclear cAMP led to a significant reduction in proliferation while a cytoplasmic sponge had little to no effect. As with the nuclear localized sponge, pharmacologically inhibiting sAC signaling suppressed astrocyte proliferation in a concentration-dependent manner, suggesting it is the source of cAMP in the nucleus. Moreover, sAC inhibition led to cell cycle arrest at the G1-S phase transition, implying that sAC-derived cAMP maybe required to enter S-phase. These results were also confirmed in primary sAC knockout astrocytes, supporting this mechanism.

Conclusions : Here we describe a novel mechanism by which compartmentalized cAMP in the nucleus regulates astrocyte proliferation. Our data suggests that sAC is the source of relevant cAMP during cell cycle progression required to transition from G1 to S-phase. In the future, understanding how astrocyte proliferation in vivo is affected by sAC signaling will add significance to these data in a broader context.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.


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