Daniel R. Storm

Cross Talk between ERK and PKA Is Required for Ca2+ Stimulation of CREB-Dependent Transcription and ERK Nuclear Translocation

Although Ca2+-stimulated cAMP response element binding protein- (CREB-) dependent transcription has been implicated in growth, differentiation, and neuroplasticity, mechanisms for Ca2+-activated transcription have not been defined. Here, we report that extracellular signal-related protein kinase (ERK) signaling is obligatory for Ca2+-stimulated transcription in PC12 cells and hippocampal neurons. The sequential activation of ERK and Rsk2 by Ca2+ leads to the phosphorylation and transactivation of CREB. Interestingly, the Ca2+-induced nuclear translocation of ERK and Rsk2 to the nucleus requires protein kinase A (PKA) activation. This may explain why PKA activity is required for Ca2+-stimulated CREB-dependent transcription. Furthermore, the full expression of the late phase of long-term potentiation (L-LTP) and L-LTP-associated CRE-mediated transcription requires ERK activation, suggesting that the activation of CREB by ERK plays a critical role in the formation of long lasting neuronal plasticity.

Induction of CRE-mediated gene expression by stimuli that generate long-lasting LTP in area CA1 of the hippocampus.

Gene expression regulated by the cAMP response element (CRE) has been implicated in synaptic plasticity and long-term memory. It has been proposed that CRE-mediated gene expression is stimulated by signals that induce long-term potentiation (LTP). To test this hypothesis, we made mice transgenic for a CRE-regulated reporter construct. We focused on long-lasting long-term potentiation (L-LTP), because it depends on cAMP-dependent protein kinase activity (PKA) and de novo gene expression. CRE-mediated gene expression was markedly increased after L-LTP, but not after decremental UP (D-LTP). Furthermore, inhibitors of PKA blocked L-LTP and associated increases in CRE-mediated gene expression. These data demonstrate that the signaling required for the generation of L-LTP but not D-LTP is sufficient to stimulate CRE-mediated transcription in the hippocampus.

Making New Connections: Role of ERK/MAP Kinase Signaling in Neuronal Plasticity

The robust translocation of MAPK during synaptic plasticity (11xMartin, K.C, Michael, D, Rose, J.C, Barad, M, Casadio, A, Zhu, H, and Kandel, E.R. Neuron. 1997; 18: 899–912Abstract | Full Text | Full Text PDF | PubMed | Scopus (411)See all References, 9xImpey, S, Obrietan, K, Wong, S.T, Poser, S, Yano, S, Wayman, G, Deloulme, J.C, Chan, G, and Storm, D.R. Neuron. 1998; 21: 869–883Abstract | Full Text | Full Text PDF | PubMed | Scopus (644)See all References, 15xSgambato, V, Pages, C, Rogard, M, Besson, M.J, and Caboche, J. J. Neurosci. 1998; 18: 8814–8825PubMedSee all References) indicates that there are likely additional nuclear targets of MAPK signaling other than CREB. For example, several recent reports suggest that the transcription factor Elk1 is a major nuclear target of MAPK during synaptic plasticity and memory consolidation (2xBerman, D.E, Hazvi, S, Rosenblum, K, Seger, R, and Dudai, Y. J. Neurosci. 1998; 18: 10037–10044PubMedSee all References, 15xSgambato, V, Pages, C, Rogard, M, Besson, M.J, and Caboche, J. J. Neurosci. 1998; 18: 8814–8825PubMedSee all References).The prominent dendritic localization of activated MAPK following synaptic activity (Impey et al. 1998xImpey, S, Obrietan, K, Wong, S.T, Poser, S, Yano, S, Wayman, G, Deloulme, J.C, Chan, G, and Storm, D.R. Neuron. 1998; 21: 869–883Abstract | Full Text | Full Text PDF | PubMed | Scopus (644)See all ReferencesImpey et al. 1998) suggests that it may also have important cytosolic targets. The best example of such a target is the Aplysia cell adhesion molecule ApCAM. MAPK activity is required for the downregulation and internalization of ApCAM, a key step in the induction of LTF. This is an important observation because the Drosophila (Fas II) and murine (NCAM) homologs of ApCAM have also been implicated in neuronal plasticity.Collectively, these studies indicate that the MAPK pathway is a fundamental component of LTM formation in invertebrates and vertebrates. Thus, the MAPK cascade joins the cAMP/PKA pathway and the CREB transcriptional pathway as an evolutionarily conserved regulator of LTM consolidation (Figure 1Figure 1). Work showing that MAPK is a major activator of plasticity-associated CREB-dependent gene expression also strongly suggests that MAPK signaling facilitates memory consolidation and L-LTP by promoting de novo CREB-regulated gene expression. There are a number of unanswered questions regarding the role of MAPK in neuronal plasticity and memory formation. Is CREB a target of Ras/MAPK signaling during memory consolidation? How is MAPK activated during adaptive neuronal plasticity and memory consolidation? What are the cytosolic and nuclear targets of MAPK that facilitate memory formation and modulate synaptic efficacy? Additional research using temporally and spatially restricted transgenic technologies should help clarify and confirm the role of Ras/MAPK signaling in LTM.*To whom correspondence should be addressed (e-mail: dstorm@u.washington.edu).

Stimulation of cAMP response element (CRE)-mediated transcription during contextual learning

Recent studies suggest that the CREB-CRE transcriptional pathway is pivotal in the formation of some types of long-term memory. However, it has not been demonstrated that stimuli that induce learning and memory activate CRE-mediated gene expression. To address this issue, we used a mouse strain transgenic for a CRE-lac Z reporter to examine the effects of hippocampus-dependent learning on CRE-mediated gene expression in the brain. Training for contextual conditioning or passive avoidance led to significant increases in CRE-dependent gene expression in areas CA1 and CA3 of the hippocampus. Auditory cue fear-conditioning, which is amygdala dependent, was associated with increased CRE-mediated gene expression in the amygdala, but not the hippocampus. These data demonstrate that learning in response to behavioral conditioning activates the CRE transcriptional pathway in specific areas of brain.

Mutant mice and neuroscience: Recommendations concerning genetic background

The following scientists made significant contributions to the recommendations in this article:

Ca2+/calmodulin binds to and modulates P/Q-type calcium channels

Neurotransmitter release at many central synapses is initiated by an influx of calcium ions through P/Q-type calcium channels,, which are densely localized in nerve terminals. Because neurotransmitter release is proportional to the fourth power of calcium concentration,, regulation of its entry can profoundly influence neurotransmission. N- and P/Q-type calcium channels are inhibited by G proteins,, and recent evidence indicates feedback regulation of P/Q-type channels by calcium. Although calcium-dependent inactivation of L-type channels is well documented, little is known about how calcium modulates P/Q-type channels. Here we report a calcium-dependent interaction between calmodulin and a novel site in the carboxy-terminal domain of the α1A subunit of P/Q-type channels. In the presence of low concentrations of intracellular calcium chelators, calcium influx through P/Q-type channels enhances channel inactivation, increases recovery from inactivation and produces a long-lasting facilitation of the calcium current. These effects are prevented by overexpression of a calmodulin-binding inhibitor peptide and by deletion of the calmodulin-binding domain. Our results reveal an unexpected association of Ca2+/calmodulin with P/Q-type calcium channels that may contribute to calcium-dependent synaptic plasticity.

Calcium-stimulated adenylyl cyclase activity is critical for hippocampus-dependent long-term memory and late phase LTP.

It is hypothesized that Ca2+ stimulation of calmodulin (CaM)-activated adenylyl cyclases (AC1 or AC8) generates cAMP signals critical for late phase LTP (L-LTP) and long-term memory (LTM). However, mice lacking either AC1 or AC8 exhibit normal L-LTP and LTM. Here, we report that mice lacking both enzymes (DKO) do not exhibit L-LTP or LTM. To determine if these defects are due to a loss of cAMP increases in the hippocampus, DKO mice were unilaterally cannulated to deliver forskolin. Administration of forskolin to area CA1 before training restored normal LTM. We conclude that Ca2+-stimulated adenylyl cyclase activity is essential for L-LTP and LTM and that AC1 or AC8 can produce the necessary cAMP signal.

Protein phosphatase 1 is a molecular constraint on learning and memory

Repetition in learning is a prerequisite for the formation of accurate and long-lasting memory. Practice is most effective when widely distributed over time, rather than when closely spaced or massed. But even after efficient learning, most memories dissipate with time unless frequently used. The molecular mechanisms of these time-dependent constraints on learning and memory are unknown. Here we show that protein phosphatase 1 (PP1) determines the efficacy of learning and memory by limiting acquisition and favouring memory decline. When PP1 is genetically inhibited during learning, short intervals between training episodes are sufficient for optimal performance. The enhanced learning correlates with increased phosphorylation of cyclic AMP-dependent response element binding (CREB) protein, of Ca2+/calmodulin-dependent protein kinase II (CaMKII) and of the GluR1 subunit of the AMPA receptor; it also correlates with CREB-dependent gene expression that, in control mice, occurs only with widely distributed training. Inhibition of PP1 prolongs memory when induced after learning, suggesting that PP1 also promotes forgetting. This property may account for ageing-related cognitive decay, as old mutant animals had preserved memory. Our findings emphasize the physiological importance of PP1 as a suppressor of learning and memory, and as a potential mediator of cognitive decline during ageing.

Disruption of the Type III Adenylyl Cyclase Gene Leads to Peripheral and Behavioral Anosmia in Transgenic Mice

Cyclic nucleotide-gated ion channels in olfactory sensory neurons (OSNs) are hypothesized to play a critical role in olfaction. However, it has not been demonstrated that the cAMP signaling is required for olfactory-based behavioral responses, and the contributions of specific adenylyl cyclases to olfaction have not been defined. Here, we report the presence of adenylyl cyclases 2, 3, and 4 in olfactory cilia. To evaluate the role of AC3 in olfactory responses, we disrupted the gene for AC3 in mice. Interestingly, electroolfactogram (EOG) responses stimulated by either cAMP- or inositol 1,4,5-triphosphate- (IP3-) inducing odorants were completely ablated in AC3 mutants, despite the presence of AC2 and AC4 in olfactory cilia. Furthermore, AC3 mutants failed several olfaction-based behavioral tests, indicating that AC3 and cAMP signaling are critical for olfactory-dependent behavior.

Light and circadian rhythmicity regulate MAP kinase activation in the suprachiasmatic nuclei

Although the circadian time-keeping properties of the suprachiasmatic nuclei (SCN) require gene expression, little is known about the signal transduction pathways that initiate transcription. Here we report that a brief exposure to light during the subjective night, but not during the subjective day, activates the p44/42 mitogen-activated protein kinase (MAPK) signaling cascade in the SCN. In addition, MAPK stimulation activates CREB (cAMP response element binding protein), indicating that potential downstream transcription factors are stimulated by the MAPK pathway in the SCN. We also observed striking circadian variations in MAPK activity within the SCN, suggesting that the MAPK cascade is involved in clock rhythmicity.