Synaptic activity induces signalling to CREB without increasing global levels of cAMP in hippocampal neurons Anna Pokorska, 1 Peter Vanhoutte, 2 Fiona J. L. Arnold, Francesca Silvagno, Giles E. Hardingham 3 and Hilmar Bading 2 MRC Laboratory of Molecular Biology, Division of Neurobiology, Hills Road, Cambridge, UK Abstract Nuclear calcium signals associated with electrical activation of neurons can control the activity of the transcription factor cAMP-response element binding protein (CREB). Yet, cAMP is thought to be the key messenger that links synaptic activity to the regulation of CREB-mediated transcription. It is gener- ally assumed that synaptic activity increases the intracellular levels of cAMP; this causes activation of the cAMP-dependent protein kinase (PKA) that regulates CREB-mediated tran- scription either directly or through controlling nuclear signal- ling of the MAP kinases/extracellular signal-regulated kinases (ERK1/2) pathway. Here we show that, in hippocampal neu- rons, synaptic activity failed to increase global levels of cAMP that would be required for the cAMP-PKA system to induce nuclear events. Even near-continuous bursting of action pot- entials, giving rise to large nuclear calcium signals and robust CREB-dependent transcription, left global intracellular levels of cAMP unchanged. These results suggest that the cAMP- PKA system does not function as the transducer of synaptic signals to the nucleus. They indicate that the known inhibitory effects of blockers of PKA on gene expression and long- lasting plasticity triggered by calcium entry reflect a gating function of basal activity of PKA that renders neurons per- missive for nuclear calcium-regulated, CREB/CBP-dependent gene expression. Keywords: cAMP, CREB, nuclear calcium, signal transduc- tion, synaptic activity, transcription. J. Neurochem. (2003) 84, 447–452. The generally accepted view is that transcriptional responses associated with electrical activation of neurons are trig- gered by calcium influx through NMDA receptors and/or L-type voltage-gated calcium channels (Szekely et al. 1989; Murphy et al. 1991; Lerea et al. 1992; Bading et al. 1993; Lerea and McNamara 1993; Bading et al. 1995; Bito et al. 1996; Deisseroth et al. 1996; Impey et al. 1996; Fields et al. 1997; Impey et al. 1998). However, it is controversial which second messenger links calcium entry to genomic events, in particular to the transcription factor complex CREB/CREB- binding protein (CBP) that plays a key role in learning processes in Drosophila, Aplysia and in the mammalian brain (Milner et al. 1998). One widely believed mechanism of signal propagation involves cAMP and the cAMP-dependent protein kinase PKA (Milner et al. 1998), a classical activator of CREB/CBP activity (Mayr and Montminy 2001). The production of cAMP following calcium entry results from the stimulation of calcium/calmodulin-dependent adenylyl cyc- lases (Poser and Storm 2001); globally elevated levels of cAMP activate PKA causing its translocation to the nucleus and activation of CREB/CBP-mediated gene expression (Mayr and Montminy 2001). PKA has also been reported to regulate nuclear import of molecules of the MAP kinase (ERK1/2) signalling pathway (Impey et al. 1998); this signalling pathway is activated by calcium entry into hippocampal neurons (Bading and Greenberg 1991) and can signal to CREB (Xing et al. 1996), although it is not Received February 18, 2002; revised manuscript received June 28, 2002; accepted September 18, 2002. Address correspondence and reprint requests to Hilmar Bading, Department of Neurobiology, Interdisciplinary Center for Neurosciences (IZN), University of Heidelberg, Im Neuenheimer Feld 364, 69120 Heidelberg, Germany. E-mail: Hilmar.Bading@uni-hd.de 1 The present address of Anna Pokorska is Biofocus Science Park, Milton Road, Cambridge, CB4 0FG, England. 2 The present address of Peter Vanhoutte and Hilmar Bading is Depart- ment of Neurobiology, Interdisciplinary Center for Neurosciences (IZN),University of Heidelberg, Im Neuenheimer Feld 364, 69120 Heidelberg, Germany. 3 The present address of Giles E. Hardingham is Department of Pre- clinical Veterinary Sciences, Royal (Dick) School of Veterinary Studies, Edinburgh University, Edinburgh EH9 1QH, UK. Abbreviations used: CBP, CREB-binding protein; CREB, cAMP- response element binding protein; ERK1/2, extracellular signal-regulated kinases; PKA, cAMP-dependent protein kinase. Journal of Neurochemistry , 2003, 84, 447–452 Ó 2003 International Society for Neurochemistry, Journal of Neurochemistry , 84, 447–452 447