Ca2ϩ /calmodulin (CaM)-dependent protein kinase II (CaMKII) "autonomy" (T286-autophosphorylation-induced Ca 2ϩ -independent activity) is required for long-term potentiation (LTP) and for learning and memory, as demonstrated by CaMKII T286A mutant mice. The Ͼ20-year-old hypothesis that CaMKII stimulation is required for LTP induction, while CaMKII autonomy is required for LTP maintenance was recently supported using the cell-penetrating fusion-peptide inhibitor antCN27. However, we demonstrate here that ant/ penetratin fusion to CN27 compromised CaMKII-selectivity, by enhancing a previously unnoticed direct binding of CaM to ant/ penetratin. In contrast to antCN27, the improved cell-penetrating inhibitor tatCN21 (5 M) showed neither CaM binding nor inhibition of basal synaptic transmission. In vitro, tatCN21 inhibited stimulated and autonomous CaMKII activity with equal potency. In rat hippocampal slices, tatCN21 inhibited LTP induction, but not LTP maintenance. Correspondingly, tatCN21 also inhibited learning, but not memory storage or retrieval in a mouse in vivo model. Thus, CaMKII autonomy provides a short-term molecular memory that is important in the signal computation leading to memory formation, but is not required as long-term memory store.

A hallmark feature of Ca 2؉ /calmodulin (CaM)-dependent protein kinase II (CaMKII) regulation is the generation of Ca 2؉ -independent autonomous activity by Thr-286 autophosphorylation. CaMKII autonomy has been regarded a form of molecular memory and is indeed important in neuronal plasticity and learning/memory. Thr-286-phosphorylated CaMKII is thought to be essentially fully active (ϳ70 -100%), implicating that it is no longer regulated and that its dramatically increased Ca 2؉ / CaM affinity is of minor functional importance. However, this study shows that autonomy greater than 15-25% was the exception, not the rule, and required a special mechanism (T-site binding; by the T-substrates AC2 or NR2B). Autonomous activity toward regular R-substrates (including tyrosine hydroxylase and GluR1) was significantly further stimulated by Ca 2؉ /CaM, both in vitro and within cells. Altered K m and V max made autonomy also substrate-(and ATP) concentration-dependent, but only over a narrow range, with remarkable stability at physiological concentrations. Such regulation still allows molecular memory of previous Ca 2؉ signals, but prevents complete uncoupling from subsequent cellular stimulation. Ca 2ϩ/calmodulin (CaM) 2 -dependent protein kinase II (CaMKII) can phosphorylate a large variety of substrate proteins and is a key player in many Ca 2ϩ -regulated cellular events (for review see Refs. 1-4). However, CaMKII is best know for its regulation of long term potentiation of synaptic strength (LTP) (5, 6), likely by increasing both number (7) and single channel conductance (8, 9) of synaptic AMPA-type glutamate receptors, and possibly by stimulating BDNF production (10, 11) (for review see Refs. 1-4). CaMKII autophosphorylation at Thr-286 generates Ca 2ϩ -independent autonomous activity (12-14), a process regarded as molecular memory (for review see Ref.2) and indeed important in learning and memory (15).Phosphorylation in the activation loop is a necessary step to generate full activity of many kinases, including PKA, PKC, and several CaMKs (for review see Refs. 16,17). By contrast, CaMKII is thought to be fully activated by Ca 2ϩ /CaM alone, without requirement for phosphorylation. Its Thr-286 is not located in the activation loop, but in the N-terminal half of the autoinhibitory ␣-helix, which binds to the T-site (Thr-286-interaction site; Ref. 18) in the basal state of CaMKII (19) (Fig. 1A). The C-terminal portion of the autoinhibitory ␣-helix extends to block the substrate binding site (S-site)(19) (Fig. 1A). Ca 2ϩ /CaM binding to the autoinhibitory ␣-helix relieves the S-site block, and makes Thr-286 accessible for phosphorylation by a neighboring kinase subunit within the 12meric CaMKII holoenzyme (20 -22). Phospho-T286 then prevents complete re-binding of the autoinhibitory ␣-helix.The dual role of Ca 2ϩ /CaM in Thr-286 autophosphorylation (for kinase activation and substrate presentation) allows computation of temporal patterns in Ca 2ϩ signaling, and indeed, CaMKII autonomy is dependent on the frequency of sti...

There is increasing evidence that synapse function depends on interactions with glial cells, namely astrocytes. Studies on specific neurons of the central nervous system (CNS) indicated that glial signals also control synapse development, but it remained unclear whether this is a general principle that applies to other neuronal cell types. To address this question, we developed new methods to immunoisolate neurons from different brain regions of postnatal mice and to culture them in a chemically defined medium. Electrophysiological recordings and immunocytochemical staining revealed vigorous synaptogenesis in hippocampal and cerebellar neurons, but not in retinal ganglion cells (RGCs) in the absence of glial cells. Co-culture with glia promoted synapse formation in RGCs as indicated by a strong increase in the incidence and frequency of action potential-independent miniature synaptic currents, but showed no such effects in hippocampal or cerebellar neurons. On the other hand, glial signals promoted the efficacy of excitatory synapses in all regions as indicated by an increase in the size of spontaneous synaptic events in cerebellar cultures and of miniature synaptic currents in hippocampal neurons and RGCs. Inhibitory synaptic currents remained largely unaffected by glia. Our results indicate that in the mammalian CNS, the way that glial signals promote the development of excitatory synapses depends on the type of neuron.

Context Data and best practice recommendations for transcranial magnetic stimulation (TMS) use in adults is largely available. While there is less data in pediatric populations and no published guidelines, its practice in children continues to grow. Methods We performed a literature search through PubMed to review all TMS studies from 1985-2016 involving children and documented any adverse events. Crude risks were calculated per session. Results Following data screening, we identified 42 single pulse (spTMS) and/or paired pulse (ppTMS) TMS studies involving 639 healthy children (HC), 482 children with CNS disorders, and 84 epileptic children (EP). Adverse events (AEs) occurred at rates of 3.42%, 5.97%, and 4.55% respective to population and number of sessions. We also report 23 repetitive TMS (rTMS) studies involving 230 CNS and 24 EP with AE rates of 3.78% and 0.0% respectively. We finally identified three theta-burst stimulation (TBS) studies involving 90 HC, 40 CNS and no EP, with AE rates of 9.78% and 10.11% respectively. Three seizures were found to have occurred in CNS individuals during rTMS, with a risk of 0.14% per session. There was no significant difference in frequency of AEs by group (p = .988) nor modality (p = .928). Conclusions Available data suggests that risk from TMS/TBS in children is similar to adults. We recommend that TMS users in this population follow the most recent adult safety guidelines until sufficient data are available for pediatric specific guidelines. We also encourage continued surveillance through surveys and assessments on a session-basis.