Deconstructing Insight: EEG Correlates of Insightful Problem Solving

Background: Cognitive insight phenomenon lies at the core of numerous discoveries. Behavioral research indicates four salient features of insightful problem solving: (i) mental impasse, followed by (ii) restructuring of the problem representation, which leads to (iii) a deeper understanding of the problem, and finally culminates in (iv) an “Aha!” feeling of suddenness and obviousness of the solution. However, until now no efforts have been made to investigate the neural mechanisms of these constituent features of insight in a unified framework. Methodology/Principal Findings: In an electroencephalographic study using verbal remote associate problems, we identified neural correlates of these four features of insightful problem solving. Hints were provided for unsolved problems or after mental impasse. Subjective ratings of the restructuring process and the feeling of suddenness were obtained on trial-by-trial basis. A negative correlation was found between these two ratings indicating that sudden insightful solutions, where restructuring is a key feature, involve automatic, subconscious recombination of information. Electroencephalogram signals were analyzed in the space×time×frequency domain with a nonparametric cluster randomization test. First, we found strong gamma band responses at parieto-occipital regions which we interpreted as (i) an adjustment of selective attention (leading to a mental impasse or to a correct solution depending on the gamma band power level) and (ii) encoding and retrieval processes for the emergence of spontaneous new solutions. Secondly, we observed an increased upper alpha band response in right temporal regions (suggesting active suppression of weakly activated solution relevant information) for initially unsuccessful trials that after hint presentation led to a correct solution. Finally, for trials with high restructuring, decreased alpha power (suggesting greater cortical excitation) was observed in right prefrontal area. Conclusions/Significance: Our results provide a first account of cognitive insight by dissociating its constituent components and potential neural correlates

Validation of the specificity of the mAb 2E4E4 against CD40 expressed in swine, bovine, ovine, and caprine spleen.

<p>Immunohistochemistry performed on: A) swine; B) bovine; C) ovine; and D) caprine spleen tissues probed with the mAb 2E4E4. Background reactivity was tested by probing E) swine, F) bovine, G) ovine, and H) caprine spleen tissues with an IgG1 isotype control mAb.</p

Characterization of a Broadly Reactive Anti-CD40 Agonistic Monoclonal Antibody for Potential Use as an Adjuvant

<div><p>Lack of safe and effective adjuvants is a major hindrance to the development of efficacious vaccines. Signaling via CD40 pathway leads to enhanced antigen processing and presentation, nitric oxide expression, pro-inflammatory cytokine expression by antigen presenting cells, and stimulation of B-cells to undergo somatic hypermutation, immunoglobulin class switching, and proliferation. Agonistic anti-CD40 antibodies have shown promising adjuvant qualities in human and mouse vaccine studies. An anti-CD40 monoclonal antibody (mAb), designated 2E4E4, was identified and shown to have strong agonistic effects on primary cells from multiple livestock species. The mAb recognize swine, bovine, caprine, and ovine CD40, and evoked 25-fold or greater proliferation of peripheral blood mononuclear cells (PBMCs) from these species relative to cells incubated with an isotype control (p<0.001). In addition, the mAb induced significant nitric oxide (p<0.0001) release by bovine macrophages. Furthermore, the mAb upregulated the expression of MHC-II by PBMCs, and stimulated significant (p<0.0001) IL-1α, IL6, IL-8, and TNF-α expression by PBMCs. These results suggest that the mAb 2E4E4 can target and stimulate cells from multiple livestock species and thus, it is a potential candidate for adjuvant development. This is the first study to report an anti-swine CD40 agonistic mAb that is also broadly reactive against multiple species.</p></div

MHC-II upregulation by swine PBMCs stimulated with the mAb 2E4E4.

<p>MHC-II upregulation by swine PBMCs stimulated with the mAb 2E4E4.</p

Validation of the specificity of the mAb 2E4E4 against CD40 expressed in swine, bovine, ovine, and caprine spleen.

<p>Immunohistochemistry performed on: A) swine; B) bovine; C) ovine; and D) caprine spleen tissues probed with the mAb 2E4E4. Background reactivity was tested by probing E) swine, F) bovine, G) ovine, and H) caprine spleen tissues with an IgG1 isotype control mAb.</p

Reactivity of the mAb 2E4E4 against HEK-293A cells expressing swine or bovine CD40.

<p>Evaluation of the mAb 2E4E4 specificity against swine and bovine CD40 was performed by immunocytometric analysis: A. Analysis of mAb 2E4E4 and a defined IgG1 control mAb by PAGE; B. HEK-293A cells transfected with a construct expressing full length swine CD40 and probed with 2E4E4; C. HEK-293A cells transfected with a construct expressing full length bovine CD40 and probed with the mAb 2E4E4; and D. Sham treated HEK-293A cells probed with the mAb 2E4E4. Flow cytometric analysis performed on: E. HEK-293A cells transfected with a construct encoding full length swine CD40; or F. full length bovine CD40 probed with either the mAb 2E4E4 (Red) or IgG1 isotype control (Blue).</p

Stimulation of swine and bovine PBMC proliferation by the mAb 2E4E4.

<p>Agonistic activity of the mAb 2E4E4 on swine, bovine, ovine, and caprine PBMCs was evaluated by ³H-Thymidine incorporation. Panels: A) swine, B) bovine, C) ovine, and D) caprine PBMCs responses after incubation with 2E4E4 (representative data is shown for 3 animals from each species) or an IgG1 isotype control. Each point represents the mean SI from triplicate wells ± SD; *Significant (p<0.001) mAb 2E4E4-induced proliferation of PBMCs from all the three 3 animals compared to IgG1 isotype control treatment.</p

Bovine, ovine, and caprine CD40 protein sequences have high homology to swine CD40 protein sequence.

<p>Alignment of swine, bovine, caprine, and ovine CD40 amino acid sequences. The signal sequence is shown where the consensus sequence is highlighted in green (amino acid 1–19), whereas the consensus sequence of the transmembrane domain is highlighted in red (amino acid 192–215). The percentage identity of the extracellular domains of bovine, ovine, and caprine CD40 protein sequences to that of swine is 74%, 75%, and 75%, respectively.</p

Upregulation of pro-inflammatory cytokine response by mAb 2E4E4.

<p>Intracellular cytokine staining was used to evaluate the ability of mAb 2E4E4 to stimulate upregulation of pro-inflammatory cytokines. Swine PBMCs were incubated with the mAb 2E4E4, LPS, or media alone, harvested at 12 hr. (Gray) and 24 hr. (Black), and then probed with mAbs against A) IL-1α; B) TNF-α; C) IL-6; or D) IL-8. Each column represents the mean florescent intensity of two wells ± SD. *p<0.05.</p

Stimulation of Nitric oxide production by mAb 2E4E4.

<p>Agonistic effect of 2E4E4 was verified by nitric oxide assay using bovine monocyte-derived macrophages incubated with graded amount of the mAb 2E4E4 (Black) or IgG1 isotype control mAb (Gray). Each column represents the μM of NO₂^- mean of triplicate wells stimulated with the mAb 2E4E4 at each concentrations ± SD; n = 3; * p<0.0001.</p