Fingerprint of Circulating Immunocytes as Biomarkers for the Prognosis of Brain Inflammation and Neuronal Injury after Cardiac Arrest.

Cardiac arrest is one of the most dangerous health problems in the world. Outcome prognosis is largely based on cerebral performance categories determined by neurological evaluations. Few systemic tests are currently available to predict survival to hospital discharge. Here, we present the results from the preclinical studies of cardiac arrest and resuscitation (CAR) in mice to identify signatures of circulating immune cells as blood-derived biomarkers to predict outcomes after CAR. Two flow cytometry panels for circulating blood lymphocytes and myeloid-derived cells, respectively, were designed to correlate with neuroinflammation and neuronal and dendritic losses in the selectively vulnerable regions of bilateral hippocampi. We found that CD4+CD25+ regulatory T cells, CD11b+CD11c- and CD11b+Ly6C+Ly6G+ myeloid-derived cells, and cells positive for the costimulatory molecules CD80 and CD86 in the blood were correlated with activation of microglia and astrocytosis, and CD4+CD25+ T cells are additionally correlated with neuronal and dendritic losses. A fingerprint pattern of blood T cells and monocytes is devised as a diagnostic tool to predict CAR outcomes. Blood tests aimed at identifying these immunocyte patterns in cardiac arrest patients will guide future clinical trials to establish better prognostication tools to avoid unnecessary early withdrawal from life-sustaining treatment.

Invasion of Peripheral Immune Cells into Brain Parenchyma after Cardiac Arrest and Resuscitation.

Although a direct link has long been suspected between systemic immune responses and neuronal injuries after stroke, it is unclear which immune cells play an important role. A question remains as to whether the blood brain barrier (BBB) is transiently disrupted after circulatory arrest to allow peripheral immune cells to enter brain parenchyma. Here, we developed a clinically relevant cardiac arrest and resuscitation model in mice to investigate the BBB integrity using noninvasive magnetic resonance imaging. Changes in immune signals in the brain and periphery were assayed by immunohistochemistry and flow cytometry. Quantitative variance maps from T1-weighted difference images before and after blood-pool contrast clearance revealed BBB disruptions immediately after resuscitation and one day after reperfusion. Time profiles of hippocampal CA1 neuronal injuries correlated with the morphological changes of microglia activation. Cytotoxic T cells, CD11b+CD11c+ dendritic cells, and CD11b+CD45+hi monocytes and macrophages were significantly increased in the brain three days after cardiac arrest and resuscitation, suggesting direct infiltration of these cells following the BBB disruption. Importantly, these immune cell changes were coupled with a parallel increase in the same subset of immune cell populations in the bone marrow and blood. We conclude that neurovascular breakdown during the initial reperfusion phase contributes to the systemic immune cell invasion and subsequent neuropathogenesis affecting the long-term outcome after cardiac arrest and resuscitation.

The effect of the order in which episodic autobiographical memories versus autobiographical knowledge are shared on feelings of closeness.

Autobiographical memories (AMs) can be used to create and maintain closeness with others [Alea, N., & Bluck, S. (2003). Why are you telling me that? A conceptual model of the social function of autobiographical memory. Memory, 11(2), 165-178]. However, the differential effects of memory specificity are not well established. Two studies with 148 participants tested whether the order in which autobiographical knowledge (AK) and specific episodic AM (EAM) are shared affects feelings of closeness. Participants read two memories hypothetically shared by each of four strangers. The strangers first shared either AK or an EAM, and then shared either AK or an EAM. Participants were randomly assigned to read either positive or negative AMs from the strangers. Findings suggest that people feel closer to those who share positive AMs in the same way they construct memories: starting with general and moving to specific.

Effects of Propofol General Anesthesia on Olfactory Relearning.

How general anesthesia interferes with sensory processing to cause amnesia remains unclear. Here, we show that activation of a learning-associated immediate early gene in rat olfactory cortices is uninterrupted by propofol, an intravenous general anesthetic with putative actions on the inhibitory GABAA receptors. Once learned under anesthesia, a novel odor can no longer re-activate the same high-level transcription programming during subsequent conscious relearning. Behavioral tests indicate that the animals' ability to consciously relearn a pure odorant, first experienced under general anesthesia, is indeed compromised. In contrast, when a mixture of two novel odorants is first experienced under anesthesia and then relearned consciously in pairs with one of the components, the animals show a deficit in relearning only the component but not the mixture. Our results reveal a previously unknown mechanism of unconscious memory due to irreplaceable neuronal commitment under general anesthesia and support the notion that general anesthesia acts at stages beyond cellular coding to disrupt sensory integration for higher-order association.

Is sharing specific autobiographical memories a distinct form of self-disclosure?

Theories of autobiographical memory posit a social function, meaning that recollecting and sharing memories of specific discrete events creates and maintains relationship intimacy. Eight studies with 1,271 participants tested whether sharing specific autobiographical memories in conversations increases feelings of closeness among conversation partners, relative to sharing other self-related information. The first 2 studies revealed that conversations in which specific autobiographical memories were shared were also accompanied by feelings of closeness among conversation partners. The next 5 studies experimentally introduced specific autobiographical memories versus general information about the self into conversations between mostly unacquainted pairs of participants. Discussing specific autobiographical memories led to greater closeness among conversation partners than discussing nonself-related topics, but no greater closeness than discussing other, more general self-related information. In the final study unacquainted pairs in whom feelings of closeness had been experimentally induced through shared humor were more likely to discuss specific autobiographical memories than unacquainted control participant pairs. We conclude that sharing specific autobiographical memories may express more than create relationship closeness, and discuss how relationship closeness may afford sharing of specific autobiographical memories by providing common ground, a social display, or a safety signal.

Cellular registration without behavioral recall of olfactory sensory input under general anesthesia.

BACKGROUND: Previous studies suggest that sensory information is "received" but not "perceived" under general anesthesia. Whether and to what extent the brain continues to process sensory inputs in a drug-induced unconscious state remain unclear. METHODS: One hundred seven rats were randomly assigned to 12 different anesthesia and odor exposure paradigms. The immunoreactivities of the immediate early gene products c-Fos and Egr1 as neural activity markers were combined with behavioral tests to assess the integrity and relationship of cellular and behavioral responsiveness to olfactory stimuli under a surgical plane of ketamine-xylazine general anesthesia. RESULTS: The olfactory sensory processing centers could distinguish the presence or absence of experimental odorants even when animals were fully anesthetized. In the anesthetized state, the c-Fos immunoreactivity in the higher olfactory cortices revealed a difference between novel and familiar odorants similar to that seen in the awake state, suggesting that the anesthetized brain functions beyond simply receiving external stimulation. Reexposing animals to odorants previously experienced only under anesthesia resulted in c-Fos immunoreactivity, which was similar to that elicited by familiar odorants, indicating that previous registration had occurred in the anesthetized brain. Despite the "cellular memory," however, odor discrimination and forced-choice odor-recognition tests showed absence of behavioral recall of the registered sensations, except for a longer latency in odor recognition tests. CONCLUSIONS: Histologically distinguishable registration of sensory processing continues to occur at the cellular level under ketamine-xylazine general anesthesia despite the absence of behavioral recognition, consistent with the notion that general anesthesia causes disintegration of information processing without completely blocking cellular communications.

Four-alpha-helix bundle with designed anesthetic binding pockets. Part II: halothane effects on structure and dynamics.

As a model of the protein targets for volatile anesthetics, the dimeric four-alpha-helix bundle, (Aalpha(2)-L1M/L38M)(2), was designed to contain a long hydrophobic core, enclosed by four amphipathic alpha-helices, for specific anesthetic binding. The structural and dynamical analyses of (Aalpha(2)-L1M/L38M)(2) in the absence of anesthetics (another study) showed a highly dynamic antiparallel dimer with an asymmetric arrangement of the four helices and a lateral accessing pathway from the aqueous phase to the hydrophobic core. In this study, we determined the high-resolution NMR structure of (Aalpha(2)-L1M/L38M)(2) in the presence of halothane, a clinically used volatile anesthetic. The high-solution NMR structure, with a backbone root mean-square deviation of 1.72 A (2JST), and the NMR binding measurements revealed that the primary halothane binding site is located between two side-chains of W15 from each monomer, different from the initially designed anesthetic binding sites. Hydrophobic interactions with residues A44 and L18 also contribute to stabilizing the bound halothane. Whereas halothane produces minor changes in the monomer structure, the quaternary arrangement of the dimer is shifted by about half a helical turn and twists relative to each other, which leads to the closure of the lateral access pathway to the hydrophobic core. Quantitative dynamics analyses, including Modelfree analysis of the relaxation data and the Carr-Purcell-Meiboom-Gill transverse relaxation dispersion measurements, suggest that the most profound anesthetic effect is the suppression of the conformational exchange both near and remote from the binding site. Our results revealed a novel mechanism of an induced fit between anesthetic molecule and its protein target, with the direct consequence of protein dynamics changing on a global rather than a local scale. This mechanism may be universal to anesthetic action on neuronal proteins.

Four-alpha-helix bundle with designed anesthetic binding pockets. Part I: structural and dynamical analyses.

The four-alpha-helix bundle mimics the transmembrane domain of the Cys-loop receptor family believed to be the protein target for general anesthetics. Using high resolution NMR, we solved the structure (Protein Data Bank ID: 2I7U) of a prototypical dimeric four-alpha-helix bundle, (Aalpha(2)-L1M/L38M)(2,) with designed specific binding pockets for volatile anesthetics. Two monomers of the helix-turn-helix motif form an antiparallel dimer as originally designed, but the high-resolution structure exhibits an asymmetric quaternary arrangement of the four helices. The two helices from the N-terminus to the linker (helices 1 and 1') are associated with each other in the dimer by the side-chain ring stacking of F12 and W15 along the long hydrophobic core and by a nearly perfect stretch of hydrophobic interactions between the complementary pairs of L4, L11, L18, and L25, all of which are located at the heptad e position along the helix-helix dimer interface. In comparison, the axes of the two helices from the linker to the C-terminus (helices 2 and 2') are wider apart from each other, creating a lateral access pathway around K47 from the aqueous phase to the center of the designed hydrophobic core. The site of the L38M mutation, which was previously shown to increase the halothane binding affinity by approximately 3.5-fold, is not part of the hydrophobic core presumably involved in the anesthetic binding but shows an elevated transverse relaxation (R(2)) rate. Qualitative analysis of the protein dynamics by reduced spectral density mapping revealed exchange contributions to the relaxation at many residues in the helices. This observation was confirmed by the quantitative analysis using the Modelfree approach and by the NMR relaxation dispersion measurements. The NMR structures and Autodock analysis suggest that the pocket with the most favorable amphipathic property for anesthetic binding is located between the W15 side chains at the center of the dimeric hydrophobic core, with the possibility of two additional minor binding sites between the F12 and F52 ring stacks of each monomer. The high-resolution structure of the designed anesthetic-binding protein offers unprecedented atomistic details about possible sites for anesthetic-protein interactions that are essential to the understanding of molecular mechanisms of general anesthesia.