Dopamine D1R-neuron cacna1c deficiency: a new model of extinction therapy-resistant post-traumatic stress.

Post-traumatic stress disorder (PTSD) is characterized by persistent fear memory of remote traumatic events, mental re-experiencing of the trauma, long-term cognitive deficits, and PTSD-associated hippocampal dysfunction. Extinction-based therapeutic approaches acutely reduce fear. However, many patients eventually relapse to the original conditioned fear response. Thus, understanding the underlying molecular mechanisms of this condition is critical to developing new treatments for patients. Mutations in the neuropsychiatric risk gene CACNA1C, which encodes the Cav1.2 isoform of the L-type calcium channel, have been implicated in both PTSD and highly comorbid neuropsychiatric conditions, such as anxiety and depression. Here, we report that male mice with global heterozygous loss of cacna1c exhibit exacerbated contextual fear that persists at remote time points (up to 180 days after shock), despite successful acute extinction training, reminiscent of PTSD patients. Because dopamine has been implicated in contextual fear memory, and Cav1.2 is a downstream target of dopamine D1-receptor (D1R) signaling, we next generated mice with specific deletion of cacna1c from D1R-expressing neurons (D1-cacna1cKO mice). Notably, D1-cacna1cKO mice also show the same exaggerated remote contextual fear, as well as persistently elevated anxiety-like behavior and impaired spatial memory at remote time points, reminiscent of chronic anxiety in treatment-resistant PTSD. We also show that D1-cacna1cKO mice exhibit elevated death of young hippocampal neurons, and that treatment with the neuroprotective agent P7C3-A20 eradicates persistent remote fear. Augmenting survival of young hippocampal neurons may thus provide an effective therapeutic approach for promoting durable remission of PTSD, particularly in patients with CACNA1C mutations or other genetic aberrations that impair calcium signaling or disrupt the survival of young hippocampal neurons.

Cocaine- and stress-primed reinstatement of drug-associated memories elicit differential behavioral and frontostriatal circuit activity patterns via recruitment of L-type Ca2+ channels.

Cocaine-associated memories are critical drivers of relapse in cocaine-dependent individuals that can be evoked by exposure to cocaine or stress. Whether these environmental stimuli recruit similar molecular and circuit-level mechanisms to promote relapse remains largely unknown. Here, using cocaine- and stress-primed reinstatement of cocaine conditioned place preference to model drug-associated memories, we find that cocaine drives reinstatement by increasing the duration that mice spend in the previously cocaine-paired context whereas stress increases the number of entries into this context. Importantly, both forms of reinstatement require Cav1.2 L-type Ca2+ channels (LTCCs) in cells of the prelimbic cortex that project to the nucleus accumbens core (PrL→NAcC). Utilizing fiber photometry to measure circuit activity in vivo in conjunction with the LTCC blocker, isradipine, we find that LTCCs drive differential recruitment of the PrL→ NAcC pathway during cocaine- and stress-primed reinstatement. While cocaine selectively activates PrL→NAcC cells prior to entry into the cocaine-paired chamber, a measure that is predictive of duration in that chamber, stress increases persistent activity of this projection, which correlates with entries into the cocaine-paired chamber. Using projection-specific chemogenetic manipulations, we show that PrL→NAcC activity is required for both cocaine- and stress-primed reinstatement, and that activation of this projection in Cav1.2-deficient mice restores reinstatement. These data indicate that LTCCs are a common mediator of cocaine- and stress-primed reinstatement. However, they engage different patterns of behavior and PrL→NAcC projection activity depending on the environmental stimuli. These findings establish a framework to further study how different environmental experiences can drive relapse, and supports further exploration of isradipine, an FDA-approved LTCC blocker, as a potential therapeutic for the prevention of relapse in cocaine-dependent individuals.

Correction: Cocaine- and stress-primed reinstatement of drug-associated memories elicit differential behavioral and frontostriatal circuit activity patterns via recruitment of L-type Ca2+ channels.

A correction to this paper has been published and can be accessed via a link at the top of the paper.

Hyperoxaemia and hypoxaemia are associated with harm in patients with ARDS.

BACKGROUND: Oxygen therapy is routinely administered to mechanically ventilated patients. However, there remains uncertainty about the optimal oxygen titration target in patients with the acute respiratory distress syndrome (ARDS). METHODS: Prospectively identified adult patients meeting the Berlin definition of ARDS between 1st January 2014 and 13th December 2016 were analyzed. Oxygen exposure variables were collected at 6-hourly intervals. The primary exposure was the average time-weighted partial pressure of arterial oxygen (PaO2) calculated over a maximum of 7 days from meeting ARDS criteria. The primary outcome was ICU mortality. Univariable and multivariable logistic regression analyses were performed to assess the impact of exposure variables on clinical outcomes. Results are presented as odds ratio [95% confidence interval]. RESULTS: 202 patients were included in the final analysis. Overall ICU mortality was 31%. The average time-weighted PaO2 during the first 7 days of ARDS was similar between non-survivors and survivors (11.3 kPa [10.2, 12.5] (84.8 mmHg [76.5, 93.8]) vs. 11.9 kPa [10.9, 12.6] (89.3 mmHg [81.8, 94.5]); p = 0.08). In univariable and multivariable analysis, average time-weighted PaO2 demonstrated a U-shaped relationship with ICU mortality. There was a similar relationship identified with hospital mortality. CONCLUSIONS: In patients with ARDS, the predicted probability of both ICU and hospital mortality was lowest when the average time-weighted PaO2 was between 12.5 and 14 kPa (93.8-105.0 mmHg), suggesting this is a reasonable oxygenation target for clinicians to aim for.

Repeat investigation during social preference behavior is suppressed in male mice with prefrontal cortex cacna1c (Cav1.2)-deficiency through the dysregulation of neural dynamics.

Impairments in social behavior are observed in a range of neuropsychiatric disorders and several lines of evidence have demonstrated that dysfunction of the prefrontal cortex (PFC) plays a central role in social deficits. We have previously shown that loss of neuropsychiatric risk gene Cacna1c that codes for the Cav1.2 isoform of L-type calcium channels (LTCCs) in the PFC result in impaired sociability as tested using the three-chamber social approach test. In this study we aimed to further characterize the nature of the social deficit associated with a reduction in PFC Cav1.2 channels (Cav1.2PFCKO mice) by testing male mice in a range of social and nonsocial tests while examining PFC neural activity using in vivo GCaMP6s fiber photometry. We found that during the first investigation of the social and non-social stimulus in the three-chamber test, both Cav1.2PFCKO male mice and Cav1.2PFCGFP controls spent significantly more time with the social stimulus compared to a non-social object. In contrast, during repeat investigations while Cav1.2PFCWT mice continued to spend more time with the social stimulus, Cav1.2PFCKO mice spent equal amount of time with both social and non-social stimuli. Neural activity recordings paralleled social behavior with increase in PFC population activity in Cav1.2PFCWT mice during first and repeat investigations, which was predictive of social preference behavior. In Cav1.2PFCKO mice, there was an increase in PFC activity during first social investigation but not during repeat investigations. These behavioral and neural differences were not observed during a reciprocal social interaction test nor during a forced alternation novelty test. To evaluate a potential deficit in reward-related processes, we tested mice in a three-chamber test wherein the social stimulus was replaced by food. Behavioral testing revealed that both Cav1.2PFCWT and Cav1.2PFCKO mice showed a preference for food over object with significantly greater preference during repeat investigation. Interestingly, there was no increase in PFC activity when Cav1.2PFCWT or Cav1.2PFCKO first investigated the food however activity significantly increased in Cav1.2PFCWT mice during repeat investigations of the food. This was not observed in Cav1.2PFCKO mice. In summary, a reduction in Cav1.2 channels in the PFC suppresses the development of a sustained social preference in mice that is associated with lack of PFC neuronal population activity that may be related to deficits in social reward.

Scn2a severe hypomorphic mutation decreases excitatory synaptic input and causes autism-associated behaviors.

SCN2A, encoding the neuronal voltage-gated Na+ channel NaV1.2, is one of the most commonly affected loci linked to autism spectrum disorders (ASDs). Most ASD-associated mutations in SCN2A are loss-of-function mutations, but studies examining how such mutations affect neuronal function and whether Scn2a mutant mice display ASD endophenotypes have been inconsistent. We generated a protein truncation variant Scn2a mouse model (Scn2aΔ1898/+) by CRISPR that eliminates the NaV1.2 channel's distal intracellular C-terminal domain, and we analyzed the molecular and cellular consequences of this variant in a heterologous expression system, in neuronal culture, in brain slices, and in vivo. We also analyzed multiple behaviors in WT and Scn2aΔ1898/+ mice and correlated behaviors with clinical data obtained in human subjects with SCN2A variants. Expression of the NaV1.2 mutant in a heterologous expression system revealed decreased NaV1.2 channel function, and cultured pyramidal neurons isolated from Scn2aΔ1898/+ forebrain showed correspondingly reduced voltage-gated Na+ channel currents without compensation from other CNS voltage-gated Na+ channels. Na+ currents in inhibitory neurons were unaffected. Consistent with loss of voltage-gated Na+ channel currents, Scn2aΔ1898/+ pyramidal neurons displayed reduced excitability in forebrain neuronal culture and reduced excitatory synaptic input onto the pyramidal neurons in brain slices. Scn2aΔ1898/+ mice displayed several behavioral abnormalities, including abnormal social interactions that reflect behavior observed in humans with ASD and with harboring loss-of-function SCN2A variants. This model and its cellular electrophysiological characterizations provide a framework for tracing how a SCN2A loss-of-function variant leads to cellular defects that result in ASD-associated behaviors.

Contribution of D1R-expressing neurons of the dorsal dentate gyrus and Cav1.2 channels in extinction of cocaine conditioned place preference.

Cocaine-associated contextual cues can trigger relapse behavior by recruiting the hippocampus. Extinction of cocaine-associated contextual memories can reduce cocaine-seeking behavior, however the molecular mechanisms within the hippocampus that underlie contextual extinction behavior and subsequent reinstatement remain poorly understood. Here, we extend our previous findings for a role of Cav1.2 L-type Ca2+ channels in dopamine 1 receptor (D1R)-expressing cells in extinction of cocaine conditioned place preference (CPP) in adult male mice. We report that attenuated cocaine CPP extinction in mice lacking Cav1.2 channels in D1R-expressing cells (D1cre, Cav1.2fl/fl) can be rescued through chemogenetic activation of D1R-expressing cells within the dorsal dentate gyrus (dDG), but not the dorsal CA1 (dCA1). This is supported by the finding that Cav1.2 channels are required in excitatory cells of the dDG, but not in the dCA1, for cocaine CPP extinction. Examination of the role of S1928 phosphorylation of Cav1.2, a protein kinase A (PKA) site using S1928A Cav1.2 phosphomutant mice revealed no extinction deficit, likely due to homeostatic scaling up of extinction-dependent S845 GluA1 phosphorylation in the dDG. However, phosphomutant mice failed to show cocaine-primed reinstatement which can be reversed by chemogenetic manipulation of excitatory cells in the dDG during extinction training. These findings outline an essential role for the interaction between D1R, Cav1.2, and GluA1 signaling in the dDG for extinction of cocaine-associated contextual memories.

Acute respiratory distress syndrome subphenotypes and differential response to simvastatin: secondary analysis of a randomised controlled trial.

BACKGROUND: Precision medicine approaches that target patients on the basis of disease subtype have transformed treatment approaches to cancer, asthma, and other heterogeneous syndromes. Two distinct subphenotypes of acute respiratory distress syndrome (ARDS) have been identified in three US-based clinical trials, and these subphenotypes respond differently to positive end-expiratory pressure and fluid management. We aimed to investigate whether these subphenotypes exist in non-US patient populations and respond differently to pharmacotherapies. METHODS: HARP-2 was a multicentre, randomised controlled trial of simvastatin (80 mg) versus placebo done in general intensive care units (ICUs) at 40 hospitals in the UK and Ireland within 48 h of onset of ARDS. The primary outcome was ventilator-free days, and secondary outcomes included non-pulmonary organ failure-free days and mortality. In a secondary analysis of HARP-2, we applied latent class analysis to baseline data without consideration of outcomes to identify subphenotypes, and we compared clinical outcomes across subphenotypes and treatment groups. FINDINGS: 540 patients were recruited to HARP-2. One patient withdrew consent for the use of their data, so data from 539 patients were analysed. In our secondary analysis, a two-class (two subphenotype) model was an improvement over a one-class model (p<0·0001), with 353 (65%) patients in the hypoinflammatory subphenotype group and 186 (35%) in the hyperinflammatory subphenotype group. Additional classes did not improve model fit. Clinical and biological characteristics of the two subphenotypes were similar to previous studies. Patients with the hyperinflammatory subphenotype had fewer ventilator-free days (median 2 days [IQR 0-17] vs 18 [IQR 0-23]; p<0·0001), fewer non-pulmonary organ failure-free days (15 [0-25] vs 27 [21-28]; p<0·0001), and higher 28-day mortality (73 [39%] vs 59 [17%]; p<0·0001) than did those with the hypoinflammatory subphenotype. Although HARP-2 found no difference in 28-day survival between placebo and simvastatin, significantly different survival was identified across patients stratified by treatment and subphenotype (p<0·0001). Specifically, within the hyperinflammatory subphenotype, patients treated with simvastatin had significantly higher 28-day survival than did those given placebo (p=0·008). A similar pattern was observed for 90-day survival. INTERPRETATION: Two subphenotypes of ARDS were identified in the HARP-2 cohort, with distinct clinical and biological features and disparate clinical outcomes. The hyperinflammatory subphenotype had improved survival with simvastatin compared with placebo. These findings support further pursuit of predictive enrichment strategies in critical care clinical trials. FUNDING: UK Efficacy and Mechanism Evaluation Programme and National Institutes of Health.