The chemokine CXCL10 modulates excitatory activity and intracellular calcium signaling in cultured hippocampal neurons

Abstract

In this study, we provide evidence for direct modulatory effects of the chemokine, CXCL10, on the physiology of hippocampal neurons maintained in primary culture. CXCL10 elicited a rise in intracellular Ca²⁺ and enhanced both spontaneous and evoked electrical activity of hippocampal neurons. CXCL10-induced elevations in intracellular Ca²⁺ were associated with an increase in neuronal firing and an alteration in the relationship between the evoked Ca²⁺ signal and neuronal activity. The effects of CXCL10 were not accompanied by a shift in resting membrane potential (RMP) or input resistance. Expression of the CXCR3 chemokine receptor supports a direct effect of CXCL10 on hippocampal neurons.

Introduction

Chemokines—or chemotactic cytokines—are a class of small (8–14 kDa) signaling proteins involved in regulation of the immune system and whose primary role is the activation and trafficking of leukocytes to sites of infection or injury during inflammatory responses (reviewed in Baggiolini, 2001, Gerard and Rollins, 2001, Mackay, 2001). More recently, it has become evident that chemokines are expressed within the central nervous system (CNS), primarily during conditions of neuroinflammation. Upregulation or dysregulation of chemokine expression has been described in a number of CNS disorders, including human immunodeficiency virus (HIV)-associated dementia (HAD) (Sanders et al., 1998), multiple sclerosis (McManus et al., 1998, Miyagishi et al., 1995, Simpson et al., 1998), Alzheimer's disease (Ishizuka et al., 1997, Xia et al., 2000), brain tumors (Suliman et al., 1999), CNS trauma (Berman et al., 1996, Glabinski et al., 1996) and stroke (Che et al., 2001, Gourmala et al., 1999, Matsumoto et al., 1997, Wang et al., 1998). Thus, there is increasing interest in the role of chemokines and chemokine receptors in neuropathological conditions as players in the neuroinflammatory response, as intermediaries of neuroimmune interaction, and as mediators of neurotoxicity or neuroprotection.

Recently, evidence has emerged for physiological roles of chemokines and their receptors in the CNS (reviewed in Bajetto et al., 2002). For example, a number of chemokines have been shown to induce neuronal Ca²⁺ signaling, modulate neurotransmitter release, regulate synaptic plasticity, reduce voltage-dependent Ca²⁺ currents, and promote neuronal survival (Giovannelli et al., 1998, Limatola et al., 2002, Limatola et al., 2000a, Limatola et al., 2000b, Limatola et al., 2003, Meucci et al., 1998, Puma et al., 2001, Ragozzino et al., 1998, Ragozzino et al., 2002). Also, certain chemokines play significant roles in neuronal migration during CNS development (Ma et al., 1998, Zou et al., 1998). For example, selective deletion of either CXCL12 (SDF-1) or its receptor CXCR4, which have a monogamous interaction, disrupts the migration of granule neurons and leads to abnormal formation of the cerebellum. A similar role of CXCL12/CXCR4 has been implicated in the migratory processes occurring during morphogenesis of the dentate gyrus (Lu et al., 2002). Despite the recent reports of chemokine effects on neuronal activity, development, and survival, the actions of chemokines in the CNS have been relatively unexplored.

The chemokine CXCL10, previously known as interferon-γ protein-10 (IP-10), is an 8.7-kDa protein whose expression is critical in mounting the host defense against viral infection of the CNS (Liu et al., 2000). CXCL10 expression has been described in a number of neurological disorders including HIV-associated dementia (HAD) (Sanders et al., 1998), multiple sclerosis/experimental autoimmune encephalomyelitis (EAE) (Fife et al., 2001, Sorensen et al., 1999), and Alzheimer's disease (Xia et al., 2000). Moreover, elevated cerebrospinal fluid (CSF) levels of CXCL10 are highly prevalent in HIV-1 infection and strongly correlate with the severity of HIV-1 associated neurologic disorders and the extent of lymphocyte chemotaxis (Kolb et al., 1999). Reports on the CNS expression of CXCR3, the receptor for CXCL10, suggest the possibility of direct effects of CXCL10 on neurons (Coughlan et al., 2000, Xia et al., 2000) as well as other cell types. CXCL10 binds selectively to CXCR3, a guanosine 5′-triphosphate (GTP)-binding protein (G-protein) coupled receptor that has been reported to activate the ras/ERK signal transduction pathway in both vascular pericytes (Bonacchi et al., 2001) and neurons (Xia et al., 2000). However, little is known about the consequences of CXCR3 activation by CXCL10 on the physiology of neurons. Thus, in the current study we have investigated the effects of CXCL10 on two important aspects of neuronal physiology: intracellular Ca²⁺ dynamics and electrical activity. The experiments were carried out in a well-characterized in vitro model system, primary cultures of hippocampal neurons.