On the mechanism of synaptic depression induced by CaMKIIN, an endogenous inhibitor of CaMKII.

Activity-dependent synaptic plasticity underlies, at least in part, learning and memory processes. NMDA receptor (NMDAR)-dependent long-term potentiation (LTP) is a major synaptic plasticity model. During LTP induction, Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) is activated, autophosphorylated and persistently translocated to the postsynaptic density, where it binds to the NMDAR. If any of these steps is inhibited, LTP is disrupted. The endogenous CaMKII inhibitor proteins CaMKIINα,β are rapidly upregulated in specific brain regions after learning. We recently showed that transient application of peptides derived from CaMKIINα (CN peptides) persistently depresses synaptic strength and reverses LTP saturation, as it allows further LTP induction in previously saturated pathways. The treatment disrupts basal CaMKII-NMDAR interaction and decreases bound CaMKII fraction in spines. To unravel CaMKIIN function and to further understand CaMKII role in synaptic strength maintenance, here we more deeply investigated the mechanism of synaptic depression induced by CN peptides (CN-depression) in rat hippocampal slices. We showed that CN-depression does not require glutamatergic synaptic activity or Ca(2+) signaling, thus discarding unspecific triggering of activity-dependent long-term depression (LTD) in slices. Moreover, occlusion experiments revealed that CN-depression and NMDAR-LTD have different expression mechanisms. We showed that CN-depression does not involve complex metabolic pathways including protein synthesis or proteasome-mediated degradation. Remarkably, CN-depression cannot be resolved in neonate rats, for which CaMKII is mostly cytosolic and virtually absent at the postsynaptic densities. Overall, our results support a direct effect of CN peptides on synaptic CaMKII-NMDAR binding and suggest that CaMKIINα,β could be critical plasticity-related proteins that may operate as cell-wide homeostatic regulators preventing saturation of LTP mechanisms or may selectively erase LTP-induced traces in specific groups of synapses

Correlation of CN-depression with average CaMKII enrichment at synapses. A.

<p>Superimposed to antCN27-induced depression in juvenile rats (42±4%, for P18–P25, n = 13) it is shown the average of control experiments conducted to evaluate signal stability during long-lasting experiments with solution recirculation. In controls (“ACSF”) no drug was applied but the solution was changed by fresh oxygenated ACSF to mimic the drug washout performed in test experiments (% rundown: 1±6%, n = 4) <b>B.</b> The same as in A, for neonate animals (P7–P10; % depression = 25±5%, n = 16). Note the rundown of synaptic potentials observed in younger rats (% rundown: 20±4, n = 7). If data is compared without correcting for rundown, depression is significantly lower in neonate rats (filled symbols in A, B; t-test, p = 0.008). <b>C.</b> Summary plot of percent decrease in transmission after antCN27 treatment (last 10 min) divided by the mean spontaneous decay measured at similar time in control experiments (**: p = 8×10⁻⁵, t-test). Data from 10 neonate and 11 juvenile rats.</p

Ca²⁺ is not required for the induction of CN-depression.

<p><b>A.</b> Depression induced by brief applications of antCN27 (5 µM, 10 min) in interleaved experiments conducted in regular ACSF (“antCN27”) or in a solution with no Ca²⁺ added and containing 10 mM EGTA and 10 µM thapsigargin, Tg (“antCN27+0Ca+Tg”). Application of the last solution by itself reversibly inhibited transmission (“0Ca+Tg”). <b>B.</b> Summary plot of percent depression induced by antCN27 in regular ACSF (40±2%, n = 4), in 0-Ca⁺² solution without Tg (“0Ca”; 56±3%, n = 4; not shown in A) and in 0Ca+Tg solution (69±3%, n = 6). *, p = 0,02; **, p = 0.009; one-way ANOVA, post hoc Tukey HSD test. Data for application of 0Ca+Tg solution only is also shown (6±5%, n = 6. No significant difference with ACSF solution change experiments; p = 0.49, t-test). <b>C.</b> Blockade of NMDAR-dependent Ca²⁺ influx does not reproduce the effect of removing Ca⁺², as revealed by an independent set of interleaved experiments of brief applications of antCN27 alone or in the presence of 100 µM APV during the whole experiment (n = 4, each) <b>D.</b> Percent depression for the experiments shown in C (40±3%, for APV; 38±8%, for regular ACSF, p = 0.98, t-test).</p

Occlusion experiments for NMDAR-LTD and CN-depression (I).

<p><b>A.</b> LTD was induced by bath application of NMDA (20 µM, 5 min) and CN-depression was subsequently induced (5 µM, 30 min; n = 8). The data were realigned according to the time of second drug application. <b>B.</b> Similar experiments to A, with drugs applied in reverse order (n = 7). <b>C.</b> Percent depression caused by antCN27 applied after LTD (“2nd”; from A) is not significantly different to that observed in regular conditions (“1st”; from B) (57±4%, 1st; 24±14%, 2nd; measured 50 min after antCN27 removal; t-test, p = 0.06). <b>D.</b> The magnitude of LTD induced in regular conditions (“1st”; from A) and after CN-depression (“2nd”; from B), also shows no statistical difference (55±4%, 1st; 31±12%, 2nd; measured 45 min after NMDA removal; t-test, p = 0.07). To quantify depression induced by the second treatment, data were renormalized to the level of transmission before drug application.</p

Protein synthesis and degradation are not required for CN-depression.

<p><b>A.</b> Depression induced by antCN27 (5 µM, 30 min) in the presence of 20 µM anisomycin (Aniso) is comparable to that induced in regular ACSF. Aniso was applied at least 20 min before antCN27 and was maintained for 1 h after starting peptide treatment. <b>B.</b> Summary plot of percent depression for the experiments shown in A and for similar trials in the presence of the proteasome inhibitor MG 132 (10–20 µM) (52±6%, n = 9, for ACSF; 41±6%, n = 5, for Aniso; 42±8%, n = 4, for MG132; one-way ANOVA, p = 0.45).</p

CN-induced depression is independent from synaptic activity.

<p><b>A.</b> Persistent synaptic depression induced by transient bath application of antCN27 (5 µM, 30 min) (% depression = 45±8% = mean ± SEM, n = 5). Data were normalized relative to baseline values before drug application. A slight and reversible reduction in presynaptic fiber volley (FV) is observed only during drug application. <b>B.</b> In a series of interleaved experiments, antCN27 was applied in regular ACSF or in the presence of kynurenic acid (Kyn; 10 mM), antagonist of AMPA/kainate receptors (n = 4, each). The inhibition of transmission by Kyn itself was completely reversible (8±2%, n = 4; similar to control experiments with ACSF solution changes (see Fig. 6); t-test, p = 0.28). <b>C.</b> Preincubation with the broad mGluR antagonist LY341495 (20 mM, n = 5) had no effect on CN-depression. <b>D.</b> Summary plot of depression for the different conditions (58±8%, for ACSF; 65±7%, for Kyn and 56±5%, for LY341495; one-way ANOVA, p = 0.62). Insets in A, B are representative field potential waveforms (averages of eight consecutive recordings) obtained at the times indicated by numbers; calibration: 0.4 mV, 5 ms. Norm.: Normalized. Error bars represent SEM in all figures. % depression: average for the last 10 min of recording and relative to baseline transmission, in all figures.</p

Disrupted third visual pathway function in schizophrenia: Evidence from real and implied motion processing

Impaired motion perception in schizophrenia has been associated with deficits in social-cognitive processes and with reduced activation of visual sensory regions, including the middle temporal area (MT+) and posterior superior temporal sulcus (pSTS). These findings are consistent with the recent proposal of the existence of a specific ‘third visual pathway’ specialized for social perception in which motion is a fundamental component. The third visual pathway transmits visual information from early sensory visual processing areas to the STS, with MT+ acting as a critical intermediary. We used functional magnetic resonance imaging to investigate functioning of this pathway during processing of naturalistic videos with explicit (real) motion and static images with implied motion cues. These measures were related to face emotion recognition and motion-perception, as measured behaviorally. Participants were 28 individuals with schizophrenia (Sz) and 20 neurotypical controls. Compared to controls, individuals with Sz showed reduced activation of third visual pathway regions (MT+, pSTS) in response to both real- and implied-motion stimuli. Dysfunction of early visual cortex and pulvinar were also associated with aberrant real-motion processing. Implied-motion stimuli additionally engaged a wide network of brain areas including parietal, motor and frontal nodes of the human mirror neuron system. The findings support concepts of MT+ as a mediator between visual sensory areas and higher-order brain and argue for greater focus on MT+ contributions to social-cognitive processing, in addition to its well-documented role in visual motion processing

Different Faces of Idiopathic Pulmonary Fibrosis With Preserved Forced Vital Capacity.

Idiopathic pulmonary fibrosis (IPF) is progressive and irreversible. Some discrepancies about IPF staging exists, especially in mild phases. Forced vital capacity (FVC) higher than 80% has been considered early or mild IPF even for the design of clinical trials. Spanish multicentre, observational, retrospective study of IPF patients diagnosed between 2012 and 2016, based on the ATS/ERS criteria, which presented FVC greater or equal 80% at diagnosis. Clinical and demographic characteristics, lung function, radiological pattern, treatment, and follow-up were analyzed. 225 IPF patients were included, 72.9% were men. The mean age was 69.5 years. The predominant high-resolution computed tomography (HRCT) pattern was consistent usual interstitial pneumonia (UIP) (51.6%). 84.7% of patients presented respiratory symptoms (exertional dyspnea and/or cough) and 33.33% showed oxygen desaturation below 90% in the 6min walking test (6MWT). Anti-fibrotic treatment was initiated at diagnosis in 55.11% of patients. Median FVC was 89.6% (IQR 17) and 58.7% of patients had a decrease of diffusion lung capacity for carbon monoxide (DLCO) below 60% of theoretical value; most of them presented functional progression (61.4%) and higher mortality at 3 years (20.45%). A statistically significant correlation with the 3-years mortality was observed between DLCO Patients with preserved FVC but presenting UIP radiological pattern and moderate-severe DLCO decrease at diagnosis associate an increased risk of progression, death or lung transplantation. Therefore, in these cases, preserved FVC would not be representative of early or mild IPF