BRAIN RESEARCH ELSEVIER Brain Research 674 (1995) 314-328 Research report The role of extracellular ionic changes in upregulating the mRNA for glial fibrillary acidic protein following spreading depression Daniel J. Bonthius a,b,l Eric W. Lothman c, Oswald Steward a,. a Department of Neuroscience, University of Virginia Health Sciences Center, Charlottesville, VA 22908, USA b Department of Pediatrics, University of Virginia Health Sciences Center, Charlottesville, VA 22908, USA c Department of Neurology, University of Virginia Health Sciences Center, Charlottesville, VA 22908, USA Accepted 13 December 1994 Abstract While spreading depression has been shown to be a powerful stimulus in upregulating glial fibrillary acidic protein (GFAP) mRNA expression, the specific physiological signal underlying the upregulation is unknown. During spreading depression, extracellular ionic concentrations are altered markedly. The present study evaluates the role of these changes in extracellular ionic concentrations as potential signals influencing GFAP mRNA expression. Gel foam pledgets saturated with artificial cerebrospinal fluid (CSF) solutions in which [Na+], [Ca2+], [K +] and [H +] were altered one at a time to match concentrations seen in spreading depression were applied to exposed parietal cortex for one hour. Dot blot and in situ hybridization techniques were used to evaluate GFAP mRNA levels. We found that CSF containing 60 mM KC1 produced a dramatic upregulation of GFAP mRNA levels throughout the cerebral cortex of the ipsilateral hemisphere without causing detectable tissue damage. The pattern and time course of the change were similar to those following application of 3 M KC1. Alteration of other ionic species did not affect GFAP mRNA levels. However, the upregulation of GFAP mRNA was not likely due directly to the increased [K+], but rather to the spreading depression that the elevated [K +] induced. This was demonstrated by the finding that the upregulation in GFAP mRNA induced by the potassium exposure was totally blocked by prior administration of MK-801, an NMDA antagonist that blocks spreading depression. These results demonstrate that an upregulation in GFAP mRNA can occur in the absence of degeneration debris and that the initiating events can be related to physiological changes, but that changes in extracellular ionic concentrations are not the likely molecular signals underlying the upregulation. Keywords: Glial fibrillary acidic protein; mRNA; Spreading depression; Potassium; MK-801 1. Introduction Following a variety of injuries to the central nervous system, astrocytes are transformed from a normal to a reactive state [3,33,35,43]. These reactive astrocytes then play important roles in the response of the CNS to the injury, including scar formation [39] and removal of degenerating synaptic terminals [35]. The histologi- cal hallmarks of this transformation are hypertrophy and hyperplasia of the involved astrocytes [16], while * Corresponding author. Department of Neuroscience, University of Virginia Medical Center, Box # 5148, MR4 Annex, Lane Rd, Ext., Charlottesville, VA 22908, USA. Fax: (1) (804) 982 4380. 1 Current Address: Departments of Neurology and Pediatrics, University of Iowa College of Medicine, Iowa City, Iowa 52242 0006-8993/95/$09.50 © 1995 Elsevier Science B.V. All rights reserved SSDI 0006-8993(95)00035-6 the biochemical hallmark is an increased astrocytic content of glial fibrillary acidic protein (GFAP) [3,46], a cytoskeletal protein unique within the CNS to astro- cytes. Until recently, degenerating tissue has been thought to be the primary signal in triggering the astrocytic response [3,33]. It has recently been shown, however, that other types of signals, such as abnormal electro- chemical activity, may also influence GFAP gene ex- pression [6,44]. One event that appears to be a power- ful stimulus is spreading depression [5,25]. Spreading depression is a cessation of electrical activity which begins as a massive depolarization of cortical neurons and glia, leading to large increases in extracellular potassium concentration, release of neurotransmitters, and a large negative shift in DC potential [27]. Once initiated, spreading depression propogates slowly across