IL-1β differentially regulates calcium wave propagation between primary human fetal astrocytes via pathways involving P2 receptors and gap junction channels

Abstract

In mammalian astrocytes, calcium waves are transmitted between cells via both a gap junction-mediated pathway and an extracellular, P2 receptor-mediated pathway, which link the cells into a syncytium. Calcium waves in astrocytes have also been shown to evoke calcium transients in neurons, and activity in neurons can elicit calcium waves in astrocytes. In this study, we show that in primary human fetal astrocytes, the P2 receptor-mediated and gap junctionmediated pathways are differentially regulated by the cytokine IL-1␤. Confocal microscopy of astrocytes loaded with Indo-1 demonstrated that intercellular calcium wave transmission in IL-1␤-treated cultures was potentiated compared with controls. However, transmission of calcium waves via the gap junction-mediated pathway was strikingly reduced. The major component of functional gap junctions in human fetal astrocytes was demonstrated to be connexin43 (Cx43), and there was a marked reduction of junctional conductance, loss of dye coupling, loss of Cx43 protein, and down-regulation of Cx43 mRNA expression after IL-1␤ treatment of cultures. Conversely, transmission of calcium waves via the P2 receptormediated pathway was potentiated in IL-1␤-treated cultures compared with controls. This potentiation was associated with an increase in the number of cells responsive to UTP, and with a transient increase in expression of the P2Y 2 purinoceptor mRNA. Because in inf lammatory conditions of the human central nervous system IL-1␤ is produced both by resident glia and by invading cells of the immune system, our results suggest that inf lammatory events may have a significant impact on coordination of astrocytic function and on information processing in the central nervous system. Intercellular calcium signaling has been shown to occur in a variety of cells in culture and may represent a general mechanism for cell-cell communication (1). Two models have been proposed by which intercellular calcium waves may be transmitted in populations of nonexcitable cells. In one model, the increase in intracellular calcium concentration is transmitted by diffusion of inositol triphosphate (IP 3 ) and͞or calcium between adjacent cells via gap junctions composed of connexin subunits (2, 3). In the second, the calcium wave is mediated by autocrine or paracrine activity of secreted tri-and dinucleotide phosphates on P2 receptors, which may further be divided into the metabotropic P2Y receptor family and the ionotropic P2X receptor family (4). In the central nervous system (CNS), intercellular calcium waves have been observed in neurons (5), astrocytes (6, 7), and oligodendrocytes (8), and although these observations were originally made in populations of cells in culture, more recently intercellular calcium waves have also been observed in more integrated systems, for example in astrocytes in brain slice cultures (9), in neurons in cerebral cortical slices (5), and in glial cells in an intact preparation of retina (10). Intercellular calcium signaling in astrocytes has attracted par...

Modulation of connexin expression and gap junction communication in astrocytes by the gram‐positive bacterium S. aureus

“…In contrast, Cx26 levels were enhanced in response to S. aureus and PGN. Our findings demonstrating the activation-dependent decrease in astrocyte Cx43 expression are in agreement with other reports demonstrating that Cx43 levels are downregulated in astrocytes in response to proinflammatory mediators such as NO and IL-1β (Bolanos and Medina, 1996;Duffy et al, 2000;John et al, 1999).…”

“…In the experimental brain abscess model which has been developed in our laboratory using S. aureus, we have shown that IL-1β and TNF-α play critical roles in CNS pathology . Interestingly, these same proinflammatory cytokines have recently been reported by others to exert dramatic effects on glial homocellular GJC (Chanson et al, 2001;Duffy et al, 2000;Faustmann et al, 2003;John et al, 1999). Based upon this evidence, we propose that proinflammatory mediators released by S. aureus-activated astrocytes in developing brain abscesses may be capable of modulating the nature and extent of GJC, with changes initiated in the proximity of the lesion and consequently extending to affect the surrounding astrocytic syncytial network.…”

“…Indeed, the gram-negative bacterial cell wall component lipopolysaccharide (LPS) has been shown to attenuate GJC in primary rat astrocytes, which was attributed, in part, to the autocrine/paracrine action of nitric oxide (NO), since the iNOS inhibitor N G -monoethyl-L-arginine (NMMA) was capable of restoring gap junction coupling (Bolanos and Medina, 1996). In addition, treatment of human fetal astrocytes with the proinflammatory cytokine interleukin-1β (IL-1β) resulted in reduced Cx43 expression concomitant with an attenuation in GJC (John et al, 1999). Taken together, these findings suggest that inflammatory stimuli and proinflammatory mediators can regulate the expression of gap junction proteins and subsequent communication in astrocytes.…”

Glia: the fulcrum of brain diseases

“…For instance, exposure of cultured astrocytes to interleukin 1b (IL-1b) downregulates their content of Cx43 at both mRNA and protein levels. 42 This effect is potentiated by another proinflammatory cytokine tumour necrosis factor-a (TNF-a) as well as by b-amyloid. These inhibitory effects are reproduced by co-culturing astrocytes with activated microglia known to release proinflammatory cytokines, indicating that tight interaction occur between glial partners involved in reactive gliosis.…”

Extracellular Nucleotides Differentially Regulate Interleukin-1β Signaling in Primary Human Astrocytes: Implications for Inflammatory Gene Expression

“…Astrocytes function to maintain the homeostatic environment of the CNS and also play an important role in immune regulation, acting as a source of chemokines, cytokines, and effector molecules (Ransom and Sontheimer, 1992;Norenberg, 1997). Coordination of function in astrocyte populations is believed to occur via at least two different mechanisms: an intercellular pathway mediated by gap junctions composed of connexin43 subunits and an extracellular pathway mediated by receptors of the P2 family that respond to nucleotides such as ATP, ADP, UTP, and UDP (Guthrie et al, 1999;John et al, 1999;Scemes et al, 2000). These pathways are thought to provide a mechanism whereby astrocytes are able to sense and respond to changes in the state and activity of neighboring cells and the surrounding CNS microenvironment.…”