Nucleotide-mediated intracellular and intercellular communication in spinal astrocytes.
Published: 2004
Language: eng
Number of pages: 308
Description: The work presented in this thesis is focused on the effects of extracellular nucleotides on dorsal spinal astrocytes. Extracellular nucleotides stimulate the elevation of intracellular Ca2+ in astrocytes. Ca 2+ signalling in astrocytes is a principal means for responding to environmental stimuli and increases in [Ca2+]i lead to growth, differentiation and release of neuroactive mediators. In addition, astrocytes are capable of homologous and heterologous intercellular signaling via the spread of Ca2+ waves.Dorsal spinal astrocytes were found to spread waves of elevated intracellular calcium to other astrocytes following mechanical stimulation. The spread of Ca2+ wave was prevented in the presence of blockers of G-protein coupled nucleotide receptors. The G-protein coupled nucleotide receptors on dorsal spinal astrocytes were identified, both molecularly and pharmacologically, as the P2Y1 and P2Y2 receptors. Heterologous expression of either of these receptors in the 1321N1 cell line was sufficient to permit the spread of Ca 2+ waves between these cells. To probe for differences in signaling between these two receptors, P2Y1 or P2Y2 receptors on spinal astrocytes were exposed to continuous application of specific agonists for each receptor. Persistent activation of the P2Y2 receptor generated a prolonged elevation of [Ca2+]i which was converted to a transient elevation in the absence of extracellular Ca2+. Stimulation of P2Y1 receptors generated only a transient elevation in [Ca2+]i.These results reveal the specific role of P2Y receptors in the mechanisms underlying the spread of Ca2+ waves in spinal astrocytes and this may represent a general principle underlying gliotransmission in the CNS.The characteristics of Ca2+ waves spreading via activation of each receptor subtype were examined and a significant difference was found in the rate at which Ca2+ waves spread in cells expressing one or the other receptor. P2Y1-1321N1 cells spread Ca2+ waves slowly, whilst in P2Y2-1321N1 cells Ca2+ waves spread significantly faster. The basis of this difference was determined to be due to a difference in the rate at which P2Y1 and P2Y2 receptors generate a Ca2+ i elevation in response to stimulation by ATP. Stimulation of P2Y1 receptors with ADP, its preferred agonist, generated a rate of Ca2+ signaling comparable to that of P2Y2 receptors. This was confirmed by forcing Ca 2+ waves to propagate via ADP in cell lines expressing P2Y1 receptors only, where Ca2+ waves were observed to spread as quickly as those in P2Y2-expressing cells.
Notes: Adviser:  Michael Salter.

Thesis (Ph.D.)--University of Toronto, 2004.

Electronic version licensed for access by U. of T. users.

Source: Dissertation Abstracts International, Volume: 65-05, Section: B, page: 2271.