RESUMEN
Calcium (Ca(2+))/calmodulin (CaM)-dependent kinase II (CaMKII) activity plays a fundamental role in learning and memory. A key feature of CaMKII in memory formation is its ability to be regulated by autophosphorylation, which switches its activity on and off during synaptic plasticity. The synaptic scaffolding protein CASK (calcium (Ca(2+))/calmodulin (CaM) associated serine kinase) is also important for learning and memory, as mutations in CASK result in intellectual disability and neurological defects in humans. We show that in Drosophila larvae, CASK interacts with CaMKII to control neuronal growth and calcium signaling. Furthermore, deletion of the CaMK-like and L27 domains of CASK (CASK ß null) or expression of overactive CaMKII (T287D) produced similar effects on synaptic growth and Ca(2+) signaling. CASK overexpression rescues the effects of CaMKII overactivity, consistent with the notion that CASK and CaMKII act in a common pathway that controls these neuronal processes. The reduction in Ca(2+) signaling observed in the CASK ß null mutant caused a decrease in vesicle trafficking at synapses. In addition, the decrease in Ca(2+) signaling in CASK mutants was associated with an increase in Ether-à-go-go (EAG) potassium (K(+)) channel localization to synapses. Reducing EAG restored the decrease in Ca(2+) signaling observed in CASK mutants to the level of wildtype, suggesting that CASK regulates Ca(2+) signaling via EAG. CASK knockdown reduced both appetitive associative learning and odor evoked Ca(2+) responses in Drosophila mushroom bodies, which are the learning centers of Drosophila. Expression of human CASK in Drosophila rescued the effect of CASK deletion on the activity state of CaMKII, suggesting that human CASK may also regulate CaMKII autophosphorylation.
RESUMEN
In humans, KCNQ2/3 channels form an M-current that regulates neuronal excitability, with mutations in these channels causing benign neonatal familial convulsions. The M-current is important in mechanisms of neural plasticity underlying associative memory and in the response to ethanol, with KCNQ controlling the release of dopamine after ethanol exposure. We show that dKCNQ is broadly expressed in the nervous system, with targeted reduction in neuronal KCNQ increasing neural excitability and KCNQ overexpression decreasing excitability and calcium signalling, consistent with KCNQ regulating the resting membrane potential and neural release as in mammalian neurons. We show that the single KCNQ channel in Drosophila (dKCNQ) has similar electrophysiological properties to neuronal KCNQ2/3, including conserved acute sensitivity to ethanol block, with the fly channel (IC(50)â=â19.8 mM) being more sensitive than its mammalian ortholog (IC(50)â=â42.1 mM). This suggests that the role of KCNQ in alcohol behaviour can be determined for the first time by using Drosophila. We present evidence that loss of KCNQ function in Drosophila increased sensitivity and tolerance to the sedative effects of ethanol. Acute activation of dopaminergic neurons by heat-activated TRP channel or KCNQ-RNAi expression produced ethanol hypersensitivity, suggesting that both act via a common mechanism involving membrane depolarisation and increased dopamine signalling leading to ethanol sedation.