Balancing risk-return decisions by manipulating the mesofrontal circuits in primates.

Decision-making is always coupled with some level of risk, with more pathological forms of risk-taking decisions manifesting as gambling disorders. In macaque monkeys trained in a high risk-high return (HH) versus low risk-low return (LL) choice task, we found that the reversible pharmacological inactivation of ventral Brodmann area 6 (area 6V) impaired the risk dependency of decision-making. Selective optogenetic activation of the mesofrontal pathway from the ventral tegmental area (VTA) to the ventral aspect of 6V resulted in stronger preference for HH, whereas activation of the pathway from the VTA to the dorsal aspect of 6V led to LL preference. Finally, computational decoding captured the modulations of behavioral preference. Our results suggest that VTA inputs to area 6V determine the decision balance between HH and LL.

How to study subjective experience in an animal model of blindsight?

The nature of subjective conscious experience, which accompanies us throughout our waking lives, and how it is generated, remain elusive. One of the challenges in studying subjective experience is disentangling the brain activity related to the sensory stimulus processing and stimulus-guided behavior from those associated with subjective perception. Blindsight, a phenomenon characterized by the retained visual discrimination performance but impaired visual consciousness due to damage to the primary visual cortex, becomes a special entry point to address this question. However, to fully understand the underlying neural mechanism, relying on studies involving human patients alone is insufficient. In this paper, we tried to address this issue, by first introducing the well-known cases of blindsight, especially the reports on subjective experience in both human and monkey subjects. And then we described how the impaired visual awareness of blindsight monkeys has been discovered and further studied by specifically designed tasks, as verbal reporting is not possible for these animals. Our previous studies also demonstrated that many complex visually guided cognitive processes were still retained despite the impairment of visual awareness. Further investigation needs to be conducted to explore the relationship between visually guided behavior, visual awareness and brain activity in blindsight subjects.

Protocol for making an animal model of "blindsight" in macaque monkeys.

Patients with damage to the primary visual cortex (V1) can respond correctly to visual stimuli in their lesion-affected visual field above the chance level, an ability named blindsight. Here, we present a protocol for making an animal model of blindsight in macaque monkeys. We describe the steps to perform pre-lesion training of monkeys on a visual task, followed by lesion surgery, post-lesion training, and evaluation of blindsight. This animal model can be used to investigate the source of visual awareness. For complete details on the use and execution of this protocol, please refer to Yoshida et al. (2008)1 and Takakuwa et al. (2021).2.

IL-23 production in human macrophages is regulated negatively by tumor necrosis factor α-induced protein 3 and positively by specificity protein 1 after stimulation of the toll-like receptor 7/8 signaling pathway.

The IL-23/IL-17 axis plays an important role in the development of autoimmune diseases, but the mechanism regulating IL-23 production is mainly unknown. We investigated how TNFAIP3 and Sp1 affect IL-23 production by human macrophages after exposure to resiquimod, a TLR7/8 agonist. IL-23 production was significantly upregulated by resiquimod but only slightly by LPS (a TLR4 agonist). Interestingly, IL-23 levels were significantly attenuated after sequential stimulation with LPS and resiquimod, but IL-12p40 and IL-18 levels were not. TLR4-related factors induced by LPS may regulate IL-23 expression via TLR7/8 signaling. LPS significantly enhanced TNFAIP3 and IRAK-M levels but reduced Sp1 levels. After exposure to resiquimod, RNA interference of TNFAIP3 upregulated IL-23 significantly more than siRNA transfection of IRAK-M did. In contrast, knockdown of Sp1 by RNA interference significantly attenuated IL-23 production. Transfection with siRNA for TNFAIP3 enhanced IL-23 expression significantly. After stimulation with resiquimod, GW7647-an agonist for PPARα (an inducer of NADHP oxidase)-and siRNA for UCP2 (a negative regulator of mitochondrial ROS generation) enhanced TNFAIP3 and reduced IL-23. siRNA for p22phox and gp91phox slightly increased Sp1 levels. However, after exposure to resiquimod siRNA-mediated knockout of DUOX1/2 significantly enhanced Sp1 and IL-23 levels, and decreased TNFα-dependent COX-2 expression. Concomitantly, TNFAIP3 levels was attenuated by DUOX1/2 siRNA. TNFAIP3 and Sp1 levels are reciprocally regulated through ROS generation. In conclusion, after stimulation of the TLR7/8 signaling pathway IL-23 production in human macrophages is regulated negatively by TNFAIP3.

TRIM28/TIF1ß and Fli-1 negatively regulate peroxynitrite generation via DUOX2 to decrease the shedding of membrane-bound fractalkine in human macrophages after exposure to substance P.

The chemokine fractalkine is synthesized as a membrane-bound protein, but studies have shown that serum levels of soluble fractalkine are elevated in inflammatory and autoimmune diseases. Patients with autoimmune diseases also have increased serum levels of neuropeptide substance P (SP). The shedding activity of the ADAM family is induced by peroxynitrite, but that of SP is unclear. Treatment of human macrophages with SP upregulated levels of membrane-bound fractalkine. Interestingly, small interfering RNA (siRNA) for DUOX2 further increased membrane-bound fractalkine but decreased soluble fractalkine compared with cells treated with SP alone. SP induced nitric oxide 2/inducible nitric oxide synthase (NOS2/iNOS) mRNA and increased levels of nitrotyrosine, a biomarker of peroxynitrite, whereas transfection with DUOX2 siRNA blunted upregulation of nitrotyrosine. Most importantly, N(ω)-nitro-L-arginine methyl ester (L-NAME, a nitric oxide synthase inhibitor) decreased protein levels of nitrotyrosine and concomitantly increased expression of membrane-bound fractalkine after exposure to SP. As for the signaling pathway of TGFß1 (an inhibitor of iNOS mRNA expression), silencing of RNA for TAK-1 upregulated membrane-bound fractalkine, but silencing of RNA for the Smad family did not. Interfering RNA of transcription factor specificity protein 1 (Sp1) upregulated protein levels of TGFß1/LAP. Most importantly, double transfection with siRNA for Sp1 and TRIM28/TIF1ßor Fli-1 led to a significant increase in TGFß1/LAP levels and a corresponding reduction of NOS2/iNOS, which inhibited the shedding of membrane-bound fractalkine. In conclusion, TRIM28/TIF1ß and Fli-1 negatively regulate TGFß1 expression to upregulate the generation of peroxynitrite, leading to increased shedding of membrane-bound fractalkine induced by SP.

An intrinsic algorithm for viewing angle tolerance of object discrimination in human subjects.

Prevailing theories suggest that view-invariant object recognition is accomplished via spatiotemporal correlations of multiple views that bind different views to the same object. However, it is unknown how the exposure to multiple views without association affects view-invariant recognition generating. Behavioural studies have shown that monkeys acquired view-invariant object recognition capability in a range of the viewing angles of 30° to 60° after experiencing discrimination of similar objects at each of several viewing angles without associating different views, but the monkeys could not discriminate novel objects from similar distractors when the viewing angle changed. In the present study the development of the view invariance was tested in the human subjects and compared with the results of the monkeys previously reported. The view-invariant object recognition capability of human subjects was tested using either familiar objects that the subjects experienced in a preparatory object discrimination task at the same viewpoints or novel objects that the subjects had never experienced. In the viewing angle range of 30°, human subjects showed significant object discrimination capability across views, with no need for prior experience of the objects. Prior discrimination experience within the same viewpoints endowed the human subjects with broadening of the viewing angle tuning, because an object discrimination test immediately after the prior discrimination experience at a single view showed wider viewing angle tuning than a test without prior experience. (PsycINFO Database Record (c) 2020 APA, all rights reserved).

Alternative routes for recovery of hand functions after corticospinal tract injury in primates and rodents.

PURPOSE OF REVIEW: Recent studies on various corticospinal tract (CST) lesions have shown the plastic changes at a variety of motor systems after the lesion. This review provides the alternative routes associated with the motor functional recovery after the CST lesions at various levels in nonhuman primates and rodents. RECENT FINDINGS: In the case of the motor cortical lesions, the perilesional area compensates for the lesion. In contrast, sprouting of the corticoreticular tracts was observed after the lesions involving sensorimotor cortical areas. After the internal capsule lesion, sprouting in the cortico-rubral pathway contributes to the recovery. In case of the pyramidal lesion, rubrospinal and reticulospinal tracts play a role of the functional recovery. After the dorsolateral funiculus (DLF) lesion at C4/C5, the indirect pathway via propriospinal tract contributes to the recovery. In case of the hemisection at lower cervical cord, the CST fibers sprouted from the bilateral motor cortex and descended to the contralesional DLF and crossed below the lesion area. SUMMARY: The central pathways can change their structure and activity dynamically depending on the lesion sites and size. Revealing the difference of the alternative pathways should be crucial to understand the whole recovery mechanism and develop the further neurorehabilitative treatment.

Di-(2-Ethylhexyl) Phthalate Promotes Release of Tissue Factor-Bearing Microparticles From Macrophages via the TGFß1/Smad/PAI-1 Signaling Pathway.

BACKGROUND: Plasminogen activator inhibitor type 1 promotes formation of endothelial microparticles with procoagulant activity. However, it remains unclear whether di-(2-ethylhexyl) phthalate, a peroxisome proliferator-activated receptor α agonist, influences microparticle formation. MATERIALS AND METHODS: The effect of di-(2-ethylhexyl) phthalate on release of tissue factor-bearing microparticles was investigated using human M1 macrophages. RESULTS: Exposure of M1 macrophages to di-(2-ethylhexyl) phthalate significantly upregulated expression of plasminogen activator inhibitor type 1, whereas incubation of macrophages with small interfering RNA for peroxisome proliferator-activated receptor α attenuated it. Di-(2-ethylhexyl) phthalate significantly increased the tissue factor protein level in culture supernatants of M1 macrophages, but not M2 macrophages. After purification of proteins by centrifugal filtration, western blotting detected 2 high molecular weight bands of tissue factor-bearing microparticles in culture supernatants of M1 macrophages. The upper band showed binding to factor VIIa and tissue factor pathway inhibitor, unlike the lower band. This suggested heterogeneity of the procoagulant activity of tissue factor-bearing microparticles, presumably dependent upon encryption/decryption of tissue factor. Phosphatidylserine contributes to tissue factor decryption, and western blotting revealed that the density of phosphatidylserine was reduced in the upper tissue factor band compared with the lower band. Di-(2-ethylhexyl) phthalate also upregulated transforming growth factor-ß1 protein production by M1 macrophages. Moreover, silencing of Smad2, Smad3 or Smad4 attenuated plasminogen activator inhibitor type 1 expression and tissue factor-release from macrophages after di-(2-ethylhexyl) phthalate stimulation. CONCLUSIONS: Di-(2-ethylhexyl) phthalate promotes formation of tissue factor-bearing microparticles in human M1 macrophages via the transforming growth factor-ß1/Smad/ plasminogen activator inhibitor type 1 signaling pathway.

Cross-talk between the transcription factor Sp1 and C/EBPß modulates TGFß1 production to negatively regulate the expression of chemokine RANTES.

RANTES is a key chemokine for atherosclerosis, and obesity is associated with progression of atherosclerosis. Substance P (SP) increases glucose uptake and accumulation of lipids in adipocytes, and SP may upregulate RANTES expression. This study investigated the mechanism of RANTES expression by human M1 macrophages stimulated with SP. SP upregulated RANTES protein expression, whereas aprepitant (an NK1R antagonist) blunted this response. Pretreatment of macrophages with BIRB796 (a combined p38γ/p38δ inhibitor) led to a significant decrease of RANTES expression. Next, we investigated the effect of several NK1R internalization factors on RANTES expression, including GRK2, ß-arrestin 2, dynamin, ROCK, and TGFß1. Exposure of macrophages to SP upregulated TGFß1 expression. Silencing of ß-arrestin 2 or GRK2 significantly enhanced the RANTES protein level after stimulation by SP, whereas TGFß1/2/3 siRNA or dynasore (a dynamin inhibitor) decreased RANTES and Y-27632 (a ROCK inhibitor) had no effect. Surprisingly, silencing of transcription factor specificity protein 1 (Sp1) or inhibition of Sp1 activity by mithramycin led to significant upregulation of TGFß1 protein and corresponding enhancement of RANTES expression (by ELISA or western blotting), whereas siRNA for C/EBPß attenuated expression of both TGFß1 and RANTES. Next, we investigated transcriptional cross-talk among Sp1 and C/EBPß, TIF1ß, or Fli-1 in relation to RANTES expression. Compared with TIF1ß or Fli-1 siRNA, C/EBPß siRNA showed significantly stronger inhibition of RANTES production by Sp1 siRNA-transfected macrophages after stimulation with SP. In conclusion, transcription factor Sp1 engages in cross-talk with C/EBPß and modulates TGFß1 production to negatively regulate RANTES expression in macrophages stimulated with SP. In conclusion, cross-talk between the transcription factor Sp1 and C/EBPß modulates TGFß1 production to negatively regulate expression of the atherogenic chemokine RANTES in SP-stimulated macrophages, while RANTES is upregulated by SP via the p38γδMAPK/C/EBPß/TGFß1 signaling pathway.

Transcription factor specificity protein 1 modulates TGFß1/Smad signaling to negatively regulate SIGIRR expression by human M1 macrophages stimulated with substance P.

The stimuli inducing expression of single immunoglobulin IL-1-related receptor (SIGIRR) and the relevant regulatory mechanisms are not well defined. Transforming growth factor ß1 (TGFß1) delays internalization of neurokinin-1 receptor (NK1R) and subsequently enhances cellular signaling. This study investigated the effect of TGFß1 on SIGIRR protein production by human M1 macrophages in response to stimulation with substance P (SP). SP caused upregulation of SIGIRR expression in a concentration-dependent manner, whereas aprepitant (an NK1R inhibitor) blunted this response. Silencing p38γMAPK or TAK-1 partially attenuated the response to SP stimulation, while TGFß1/2/3 siRNA dramatically diminished it. SP induced much greater SIGIRR protein production than either lipopolysaccharide (a TLR4 agonist) or resiquimod (a TLR7/8 agonist). Unexpectedly, silencing of transcription factor specificity protein 1 (Sp1) led to significant upregulation of SIGIRR expression after SP stimulation, while KLF2 siRNA only partially enhanced it and Fli-1 siRNA reduced it. SP also upregulated TGFß1 expression, along with a corresponding increase of SIGIRR protein, whereas silencing TGFß1/2/3 blunted these responses. Sp1 siRNA or mithramycin (a gene-selective Sp1 inhibitor) significantly enhanced the expression of TGFß1 and SIGIRR by macrophages after SP stimulation. Importantly, this effect of Sp1 siRNA on TGFß1 and SIGIRR was blunted by siRNA for Smad2, Smad3, or Smad4, but not by TAK-1 siRNA. Next, we investigated the influence of transcription factor cross-talk on SIGIRR expression in response to SP. Co-transfection of macrophages with Sp1 siRNA and C/EBPß or TIF1ß siRNA attenuated the upregulation of SIGIRR by SP, while a combination of Sp1 siRNA and Fli-1 siRNA dramatically diminished it. In conclusion, TGFß1 may be an intermediary between SP/NK1R activation and SIGIRR expression in Sp1 siRNA-transfected macrophages. In addition, Sp1 modulates TGFß1/Smad signaling and negatively regulates SIGIRR protein production by macrophages after SP stimulation.

Dynamics of population coding for object views following object discrimination training.

We have previously demonstrated that inferotemporal neurons respond to objects viewed from a range of angles, even without any prior experience in learning the associations among the views. Several models have been proposed to explain object recognition across disparate views. However, direct neuronal evidence is rare. In the present study, we focused on the response similarity of a population of inferotemporal cells to object views, following different prior experiences. Two monkeys were subjected to a task in which object discrimination across views was required. We found significantly higher neural response similarity to 30° separated views, 190ms after object image presentation, than without any prior discrimination experience across views. The time period over which the similarity was significant began and endured similarly for 60° separated views at 190-850ms. For 90° separated views, the time period over which the similarity was significant was shorter and started later, at 230-550ms. The results demonstrate the dynamics of cell population activity and suggest a possible explanation for object recognition across disparate views.

Neural substrates of view-invariant object recognition developed without experiencing rotations of the objects.

One fails to recognize an unfamiliar object across changes in viewing angle when it must be discriminated from similar distractor objects. View-invariant recognition gradually develops as the viewer repeatedly sees the objects in rotation. It is assumed that different views of each object are associated with one another while their successive appearance is experienced in rotation. However, natural experience of objects also contains ample opportunities to discriminate among objects at each of the multiple viewing angles. Our previous behavioral experiments showed that after experiencing a new set of object stimuli during a task that required only discrimination at each of four viewing angles at 30° intervals, monkeys could recognize the objects across changes in viewing angle up to 60°. By recording activities of neurons from the inferotemporal cortex after various types of preparatory experience, we here found a possible neural substrate for the monkeys' performance. For object sets that the monkeys had experienced during the task that required only discrimination at each of four viewing angles, many inferotemporal neurons showed object selectivity covering multiple views. The degree of view generalization found for these object sets was similar to that found for stimulus sets with which the monkeys had been trained to conduct view-invariant recognition. These results suggest that the experience of discriminating new objects in each of several viewing angles develops the partially view-generalized object selectivity distributed over many neurons in the inferotemporal cortex, which in turn bases the monkeys' emergent capability to discriminate the objects across changes in viewing angle.