Dopamine facilitates associative memory encoding in the entorhinal cortex.

Mounting evidence shows that dopamine in the striatum is critically involved in reward-based reinforcement learning1,2. However, it remains unclear how dopamine reward signals influence the entorhinal-hippocampal circuit, another brain network that is crucial for learning and memory3-5. Here, using cell-type-specific electrophysiological recording6, we show that dopamine signals from the ventral tegmental area and substantia nigra control the encoding of cue-reward association rules in layer 2a fan cells of the lateral entorhinal cortex (LEC). When mice learned novel olfactory cue-reward associations using a pre-learned association rule, spike representations of LEC fan cells grouped newly learned rewarded cues with a pre-learned rewarded cue, but separated them from a pre-learned unrewarded cue. Optogenetic inhibition of fan cells impaired the learning of new associations while sparing the retrieval of pre-learned memory. Using fibre photometry, we found that dopamine sends novelty-induced reward expectation signals to the LEC. Inhibition of LEC dopamine signals disrupted the associative encoding of fan cells and impaired learning performance. These results suggest that LEC fan cells represent a cognitive map of abstract task rules, and that LEC dopamine facilitates the incorporation of new memories into this map.

Vaccination with Intradermal Bacillus Calmette-Guérin Provides Robust Protection against Extrapulmonary Tuberculosis but Not Pulmonary Infection in Cynomolgus Macaques.

Recently, the efficacy of Mycobacterium bovis bacillus Calmette-Guérin (BCG) vaccination is being reassessed in accordance with the achievements of clinical tuberculosis (TB) vaccine research. However, the mechanisms ultimately determining the success or failure of BCG vaccination to prevent pulmonary TB remain poorly understood. In this study, we analyzed the protective effects of intradermal BCG vaccination by using specific pathogen-free cynomolgus macaques of Asian origin that were intradermally vaccinated with BCG (Tokyo strain) followed by Mycobacterium tuberculosis (Erdman strain) infection. Intradermal BCG administration generated TB Ag-specific multifunctional CD4 T cell responses in peripheral blood and bronchoalveolar lavage and almost completely protected against the development of TB pathogenesis with aggravation of clinical parameters and high levels of bacterial burdens in extrapulmonary organs. However, interestingly, there were no differences in bacterial quantitation and pathology of extensive granulomas in the lungs between BCG-vaccinated monkeys and control animals. These results indicated that the changes in clinical parameters, immunological responses, and quantitative gross pathology that are used routinely to determine the efficacy of TB vaccines in nonhuman primate models might not correlate with the bacterial burden and histopathological score in the lung as measured in this study.

Posterior shoulder capsule of the dominant arm is stiffer in baseball players than that in nonthrowing population.

BACKGROUND: Posterior shoulder capsule tightness is one of the factors for shoulder injuries in overhead athletes. Recent studies have shown the posterior capsule of the dominant arm to be stiffer than that of the nondominant arm in baseball players. However, whether posterior capsule tightness in the dominant arm is exclusive to overhead athletes remains unknown. This study aimed to investigate whether the posterior shoulder capsule of the dominant arm in baseball players is stiffer than that in nonthrowing population. METHODS: Fifteen male collegiate asymptomatic baseball players (baseball-player group) and fifteen male college students who did not partake in overhead sports (nonthrowing group) participated in this study. We measured the shear moduli of the middle and inferior posterior capsules, superior infraspinatus, inferior infraspinatus, teres minor, and posterior deltoid in the dominant arm by ultrasound shear wave elastography. We compared shear moduli between the two groups using an independent samples t-test and Mann-Whitney test. In addition, we investigated the correlation between the range of glenohumeral internal rotation and each shear modulus in each group using the Pearson correlation coefficient. RESULTS: The shear moduli in the baseball-player group were significantly higher than those in the nonthrowing group in both the middle posterior capsule (baseball-player group: 36.1 ± 5.6 kPa vs. nonthrowing group: 29.0 ± 8.6 kPa; P = .018) and inferior posterior capsule (37.1 ± 9.6 kPa vs. 27.9 ± 6.8 kPa; P = .002). However, no difference in the shear moduli of individual muscle groups was identified. The glenohumeral internal rotation range exhibited a statistically significant negative correlation with the shear modulus of the inferior posterior capsule in the baseball-player group (Pearson correlation coefficient = -0.586, P = .022). CONCLUSION: Our findings suggest that the posterior shoulder capsule of the dominant arm in baseball players is stiffer than that in nonthrowing population.

Acetylcholine from the nucleus basalis magnocellularis facilitates the retrieval of well-established memory.

Retrieval deficit of long-term memory is a cardinal symptom of dementia and has been proposed to associate with abnormalities in the central cholinergic system. Difficulty in the retrieval of memory is experienced by healthy individuals and not limited to patients with neurological disorders that result in forgetfulness. The difficulty of retrieving memories is associated with various factors, such as how often the event was experienced or remembered, but it is unclear how the cholinergic system plays a role in the retrieval of memory formed by a daily routine (accumulated experience). To investigate this point, we trained rats moderately (for a week) or extensively (for a month) to detect a visual cue in a two-alternative forced-choice task. First, we confirmed the well-established memory in the extensively trained group was more resistant to the retrieval problem than recently acquired memory in the moderately trained group. Next, we tested the effect of a cholinesterase inhibitor, donepezil, on the retrieval of memory after a long no-task period in extensively trained rats. Pre-administration of donepezil improved performance and reduced the latency of task initiation compared to the saline-treated group. Finally, we lesioned cholinergic neurons of the nucleus basalis magnocellularis (NBM), which project to the entire neocortex, by injecting the cholinergic toxin 192 IgG-saporin. NBM-lesioned rats showed severely impaired task initiation and performance. These abilities recovered as the trials progressed, though they never reached the level observed in rats with intact NBM. These results suggest that acetylcholine released from the NBM contributes to the retrieval of well-established memory developed by a daily routine.

SOCS1 Antagonist-Expressing Recombinant Bacillus Calmette-Guérin Enhances Antituberculosis Protection in a Mouse Model.

Suppressor of cytokine signaling 1 (SOCS1) plays a key role in the negative regulation of JAK/STAT signaling, which is involved in innate immunity and subsequent adaptive immunity. Bacillus Calmette-Guérin (BCG) induces upregulation of SOCS1 expression in host cells, which may lead to the suppression of immune responses by BCG via inhibition of the JAK/STAT signaling pathway. This might cause A reduction in the protective effect of a BCG vaccine. In the current study, we assessed the immune responses to and the protective efficacy of a recombinant BCG secreting a dominant negative mutant of the SOCS1 molecule (rBCG-SOCS1DN). C57BL/6 mice were immunized with rBCG-SOCS1DN or parental BCG Tokyo vaccine strain harboring an empty plasmid vector (rBCG-pSO). rBCG-SOCS1DN enhanced the activation of bone marrow-derived dendritic cells and the activation of T cells compared with those with rBCG-pSO. The amounts of IFN-γ, TNF-α, and IL-6 produced by splenocytes of rBCG-SOCS1DN-immunized mice were larger than those produced by splenocytes of rBCG-pSO-immunized mice. Moreover, the rBCG-SOCS1DN-immunized mice showed a substantial reduction in the number of CFU of Mycobacterium tuberculosis in the lungs and spleens compared with that in control BCG-immunized mice when the immunized mice were infected with a highly pathogenic M. tuberculosis strain by inhalation. These findings provide evidence for the possibility of rBCG-SOCS1DN being an effective M. tuberculosis vaccine with a novel concept of rBCG as a tool for immunomodulation in host cells.

Ipsilateral-Dominant Control of Limb Movements in Rodent Posterior Parietal Cortex.

It is well known that the posterior parietal cortex (PPC) and frontal motor cortices in primates preferentially control voluntary movements of contralateral limbs. The PPC of rats has been defined based on patterns of thalamic and cortical connectivity. The anatomical characteristics of this area suggest that it may be homologous to the PPC of primates. However, its functional roles in voluntary forelimb movements have not been well understood, particularly in the lateralization of motor limb representation; that is, the limb-specific activity representations for right and left forelimb movements. We examined functional spike activity of the PPC and two motor cortices, the primary motor cortex (M1) and the secondary motor cortex (M2), when head-fixed male rats performed right or left unilateral movements. Unlike primates, PPC neurons in rodents were found to preferentially represent ipsilateral forelimb movements, in contrast to the contralateral preference of M1 and M2 neurons. Consistent with these observations, optogenetic activation of PPC and motor cortices, respectively, evoked ipsilaterally and contralaterally biased forelimb movements. Finally, we examined the effects of optogenetic manipulation on task performance. PPC or M1 inhibition by optogenetic GABA release shifted the behavioral limb preference contralaterally or ipsilaterally, respectively. In addition, weak optogenetic PPC activation, which was insufficient to evoke motor responses by itself, shifted the preference ipsilaterally; although similar M1 activation showed no effects on task performance. These paradoxical observations suggest that the PPC plays evolutionarily different roles in forelimb control between primates and rodents.SIGNIFICANCE STATEMENT In rodents, the primary and secondary motor cortices (M1 and M2, respectively) are involved in voluntary movements with contralateral preference. However, it remains unclear whether and how the posterior parietal cortex (PPC) participates in controlling multiple limb movements. We recorded functional activity from these areas using a behavioral task to monitor movements of the right and left forelimbs separately. PPC neurons preferentially represented ipsilateral forelimb movements and optogenetic PPC activation evoked ipsilaterally biased forelimb movements. Optogenetic PPC inhibition via GABA release shifted the behavioral limb preference contralaterally during task performance, whereas weak optogenetic PPC activation, which was insufficient to evoke motor responses by itself, shifted the preference ipsilaterally. Our findings suggest rodent PPC contributes to ipsilaterally biased motor response and/or planning.

A spike analysis method for characterizing neurons based on phase locking and scaling to the interval between two behavioral events.

Standard analysis of neuronal functions assesses the temporal correlation between animal behaviors and neuronal activity by aligning spike trains with the timing of a specific behavioral event, e.g., visual cue. However, spike activity is often involved in information processing dependent on a relative phase between two consecutive events rather than a single event. Nevertheless, less attention has so far been paid to such temporal features of spike activity in relation to two behavioral events. Here, we propose "Phase-Scaling analysis" to simultaneously evaluate the phase locking and scaling to the interval between two events in task-related spike activity of individual neurons. This analysis method can discriminate conceptual "scaled"-type neurons from "nonscaled"-type neurons using an activity variation map that combines phase locking with scaling to the interval. Its robustness was validated by spike simulation using different spike properties. Furthermore, we applied it to analyzing actual spike data from task-related neurons in the primary visual cortex (V1), posterior parietal cortex (PPC), primary motor cortex (M1), and secondary motor cortex (M2) of behaving rats. After hierarchical clustering of all neurons using their activity variation maps, we divided them objectively into four clusters corresponding to nonscaled-type sensory and motor neurons and scaled-type neurons including sustained and ramping activities, etc. Cluster/subcluster compositions for V1 differed from those of PPC, M1, and M2. The V1 neurons showed the fastest functional activities among those areas. Our method was also applicable to determine temporal "forms" and the latency of spike activity changes. These findings demonstrate its utility for characterizing neurons.NEW & NOTEWORTHY Phase-Scaling analysis is a novel technique to unbiasedly characterize the temporal dependency of functional neuron activity on two behavioral events and objectively determine the latency and form of the activity change. This powerful analysis can uncover several classes of latently functioning neurons that have thus far been overlooked, which may participate differently in intermediate processes of a brain function. The Phase-Scaling analysis will yield profound insights into neural mechanisms for processing internal information.

Effect of blue light-emitting diode light and antioxidant potential in a somatic cell.

Light is an indispensable part of routine laboratory work in which conventional light is generally used. Light-emitting diodes (LEDs) have come to replace conventional light, and thus could be a potent target in biomedical studies. Since blue light is a major component of visible light wavelength, in this study, using a somatic cell from the African green monkey kidney, we assessed the possible consequences of the blue spectra of LED light in future animal experiments and proposed a potent mitigation against light-induced damage. COS-7 cells were exposed to blue LED light (450 nm) and the growth and deoxyribonucleic acid (DNA) damage were assessed at different exposure times. A higher suppression in cell growth and viability was observed under a longer period of blue LED light exposure. The number of apoptotic cells increased as the light exposure time was prolonged. Reactive oxygen species (ROS) generation was also elevated in accordance to the extension of light exposure time. A comparison with dark-maintained cells revealed that the upregulation of ROS by blue LED light plays a significant role in causing cellular dysfunction in DNA in a time-dependent manner. In turn, antioxidant treatment has been shown to improve cell growth and viability under blue LED light conditions. This indicates that antioxidants have potential against blue LED light-induced somatic cell damage. It is expected that this study will contribute to the understanding of the basic mechanism of somatic cell death under visible light and maximize the beneficial use of LED light in future animal experiments.

In Vivo Spiking Dynamics of Intra- and Extratelencephalic Projection Neurons in Rat Motor Cortex.

In motor cortex, 2 types of deep layer pyramidal cells send their axons to other areas: intratelencephalic (IT)-type neurons specifically project bilaterally to the cerebral cortex and striatum, whereas neurons of the extratelencephalic (ET)-type, termed conventionally pyramidal tract-type, project ipsilaterally to the thalamus and other areas. Although they have totally different synaptic and membrane potential properties in vitro, little is known about the differences between them in ongoing spiking dynamics in vivo. We identified IT-type and ET-type neurons, as well as fast-spiking-type interneurons, using novel multineuronal analysis based on optogenetically evoked spike collision along their axons in behaving/resting rats expressing channelrhodopsin-2 (Multi-Linc method). We found "postspike suppression" (~100 ms) as a characteristic of ET-type neurons in spike auto-correlograms, and it remained constant independent of behavioral conditions in functionally different ET-type neurons. Postspike suppression followed even solitary spikes, and spike bursts significantly extended its duration. We also observed relatively strong spike synchrony in pairs containing IT-type neurons. Thus, spiking dynamics in IT-type and ET-type neurons may be optimized differently for precise and coordinated motor control.

Distinct Laterality in Forelimb-Movement Representations of Rat Primary and Secondary Motor Cortical Neurons with Intratelencephalic and Pyramidal Tract Projections.

Two distinct motor areas, the primary and secondary motor cortices (M1 and M2), play crucial roles in voluntary movement in rodents. The aim of this study was to characterize the laterality in motor cortical representations of right and left forelimb movements. To achieve this goal, we developed a novel behavioral task, the Right-Left Pedal task, in which a head-restrained male rat manipulates a right or left pedal with the corresponding forelimb. This task enabled us to monitor independent movements of both forelimbs with high spatiotemporal resolution. We observed phasic movement-related neuronal activity (Go-type) and tonic hold-related activity (Hold-type) in isolated unilateral movements. In both M1 and M2, Go-type neurons exhibited bias toward contralateral preference, whereas Hold-type neurons exhibited no bias. The contralateral bias was weaker in M2 than M1. Moreover, we differentiated between intratelencephalic (IT) and pyramidal tract (PT) neurons using optogenetically evoked spike collision in rats expressing channelrhodopsin-2. Even in identified PT and IT neurons, Hold-type neurons exhibited no lateral bias. Go-type PT neurons exhibited bias toward contralateral preference, whereas IT neurons exhibited no bias. Our findings suggest a different laterality of movement representations of M1 and M2, in each of which IT neurons are involved in cooperation of bilateral movements, whereas PT neurons control contralateral movements.SIGNIFICANCE STATEMENT In rodents, the primary and secondary motor cortices (M1 and M2) are involved in voluntary movements via distinct projection neurons: intratelencephalic (IT) neurons and pyramidal tract (PT) neurons. However, it remains unclear whether the two motor cortices (M1 vs M2) and the two classes of projection neurons (IT vs PT) have different laterality of movement representations. We optogenetically identified these neurons and analyzed their functional activity using a novel behavioral task to monitor movements of the right and left forelimbs separately. We found that contralateral bias was reduced in M2 relative to M1, and in IT relative to PT neurons. Our findings suggest that the motor information processing that controls forelimb movement is coordinated by a distinct cell population.

Primary Role of Suppressor of Cytokine Signaling 1 in Mycobacterium bovis BCG Infection.

Suppressor of cytokine signaling 1 (SOCS1) is a negative regulator of JAK/STAT signaling and is induced by mycobacterial infection. To understand the major function of SOCS1 during infection, we established a novel system in which recombinant Mycobacterium bovis bacillus Calmette-Guérin expressed dominant-negative SOCS1 (rBCG-SOCS1DN) because it would not affect the function of SOCS1 in uninfected cells. When C57BL/6 mice and RAG1-/- mice were intratracheally inoculated with rBCG-SOCS1DN, the amount of rBCG-SOCS1DN in the lungs was significantly reduced compared to that in the lungs of mice inoculated with a vector control counterpart and wild-type BCG. However, these significant differences were not observed in NOS2-/- mice and RAG1-/- NOS2-/- double-knockout mice. These findings demonstrated that SOCS1 inhibits nitric oxide (NO) production to establish mycobacterial infection and that rBCG-SOCS1DN has the potential to be a powerful tool for studying the primary function of SOCS1 in mycobacterial infection.

Simian immunodeficiency virus SIVmac239 infection and simian human immunodeficiency virus SHIV89.6P infection result in progression to AIDS in cynomolgus macaques of Asian origin.

Simian immunodeficiency virus (SIV) infection models in cynomolgus macaques are important for analysis of the pathogenesis of immunodeficiency virus and for studies on the efficacy of new vaccine candidates. However, very little is known about the pathogenesis of SIV or simian human immunodeficiency virus (SHIV) in cynomolgus macaques from different Asian countries. In the present study, we analysed the infectivity and pathogenicity of CCR5-tropic SIVmac and those of dual-tropic SHIV89.6P inoculated into cynomolgus macaques in Indonesian, Malaysian or Philippine origin. The plasma viral loads in macaques infected with either SIVmac239 or SHIV89.6P were maintained at high levels. CD4+ T cell levels in macaques infected with SIVmac239 gradually decreased. All of the macaques infected with SHIV89.6P showed greatly reduced CD4+ T-cell numbers within 6 weeks of infection. Eight of the 11 macaques infected with SIVmac239 were killed due to AIDS symptoms after 2-4.5 years, while four of the five macaques infected with SHIV89.6P were killed due to AIDS symptoms after 1-3.5 years. We also analysed cynomolgus macaques infected intrarectally with repeated low, medium or high doses of SIVmac239, SIVmac251 or SHIV89.6P. Infection was confirmed by quantitative RT-PCR at more than 5000, 300 and 500 TCID50 for SIVmac239, SIVmac251 and SHIV89.6P, respectively. The present study indicates that cynomolgus macaques of Asian origin are highly susceptible to SIVmac and SHIV infection by both intravenous and mucosal routes. These models will be useful for studies on virus pathogenesis, vaccination and therapeutics against human immunodeficiency virus/AIDS.

Calcium Signaling in MR1-Dependent Antigen Presentation of Mycobacterium tuberculosis.

MR1 is a ubiquitously expressed MHC-Ib molecule that presents microbial metabolites to MR1-restricted T cells, but there are differences in the antigen presentation pathway of an intracellular microbe compared to exogenous antigen. We have shown the importance of endosomal trafficking proteins in MR1-dependent presentation of Mycobacterium tuberculosis (Mtb). Two pore channels (TPCs) are endosomal calcium channels that regulate endosomal trafficking. Due to their location on endosomes, we hypothesized that TPCs could be required for MR1-dependent presentation of antigens derived from the intracellular microbe Mtb. We found that TPCs are critical for the presentation of Mtb by MR1; inhibition of TPCs had no effect on MR1 presentation of extracellular (exogenous) antigens, HLA-B presentation, or HLA-II presentation. Finally, we found that the calcium sensitive trafficking protein Synaptotagmin 7 was also key in the presentation of Mtb by MR1. This calcium-dependent endosomal pathway is a novel mechanism by which the immune system can sample intracellular antigens.

Rat hippocampal CA1 region represents learning-related action and reward events with shorter latency than the lateral entorhinal cortex.

The hippocampus and entorhinal cortex are deeply involved in learning and memory. However, little is known how ongoing events are processed in the hippocampal-entorhinal circuit. By recording from head-fixed rats during action-reward learning, here we show that the action and reward events are represented differently in the hippocampal CA1 region and lateral entorhinal cortex (LEC). Although diverse task-related activities developed after learning in both CA1 and LEC, phasic activities related to action and reward events differed in the timing of behavioral event representation. CA1 represented action and reward events almost instantaneously, whereas the superficial and deep layers of the LEC showed a delayed representation of the same events. Interestingly, we also found that ramping activity towards spontaneous action was correlated with waiting time in both regions and exceeded that in the motor cortex. Such functional activities observed in the entorhinal-hippocampal circuits may play a crucial role for animals in utilizing ongoing information to dynamically optimize their behaviors.

Cholinergic modulation of response gain in the primary visual cortex of the macaque.

ACh modulates neuronal activity throughout the cerebral cortex, including the primary visual cortex (V1). However, a number of issues regarding this modulation remain unknown, such as the effect and its function and the receptor subtypes involved. To address these issues, we combined extracellular single-unit recordings and microiontophoretic administration of ACh and measured V1 neuronal responses to drifting sinusoidal grating stimuli in anesthetized macaque monkeys. ACh was found to have mostly facilitatory effects on the visual responses, although some cases of suppressive effects were also seen. To assess the functional role of ACh, we further examined how ACh modulates the stimulus contrast-response function, finding that the response gain increased with the facilitatory effect. The response facilitation was completely or strongly blocked by atropine (At), a muscarinic ACh receptor (mAChR) antagonist, in almost all neurons (96% of cells), whereas any residual effect after At administration was fully removed by mecamylamine, a nicotinic AChR (nAChR) antagonist, suggesting a predominant role for mAChRs in this mechanism. Furthermore, we found no laminar distribution bias for the facilitatory modulation, although the relative contribution of mAChRs was smaller in layer 4C than in other layers. The suppressive effect was blocked completely by At. These results demonstrate that ACh plays an important role in visual information processing in V1 by controlling the response gain via mAChRs across all cortical layers and via nAChRs, mainly in layer 4C.

Human lung-resident mucosal-associated invariant T cells are abundant, express antimicrobial proteins, and are cytokine responsive.

Mucosal-associated Invariant T (MAIT) cells are an innate-like T cell subset that recognize a broad array of microbial pathogens, including respiratory pathogens. Here we investigate the transcriptional profile of MAIT cells localized to the human lung, and postulate that MAIT cells may play a role in maintaining homeostasis at this mucosal barrier. Using the MR1/5-OP-RU tetramer, we identified MAIT cells and non-MAIT CD8+ T cells in lung tissue not suitable for transplant from human donors. We used RNA-sequencing of MAIT cells compared to non-MAIT CD8+ T cells to define the transcriptome of MAIT cells in the human lung. We show that, as a population, lung MAIT cells are polycytotoxic, secrete the directly antimicrobial molecule IL-26, express genes associated with persistence, and selectively express cytokine and chemokine- related molecules distinct from other lung-resident CD8+ T cells, such as interferon-γ- and IL-12- receptors. These data highlight MAIT cells' predisposition to rapid pro-inflammatory cytokine responsiveness and antimicrobial mechanisms in human lung tissue, concordant with findings of blood-derived counterparts, and support a function for MAIT cells as early sensors in the defense of respiratory barrier function.

Donor Unrestricted T Cells: Linking innate and adaptive immunity.

Donor Unrestricted T Cells (DURTs) are characterized by their use of antigen presentation molecules that are often invariant. As these cells recognize diverse mycobacterial antigens, often found in BCG, these cells have the potential to either serve as targets for vaccination, or as a means to enable the induction of traditional T and B cell immunity. Here, we will review specific DURT family members, and their relationship to BCG.

Disrupted Place Cell Remapping and Impaired Grid Cells in a Knockin Model of Alzheimer's Disease.

Patients with Alzheimer's disease (AD) suffer from spatial memory impairment and wandering behavior, but the brain circuit mechanisms causing such symptoms remain largely unclear. In healthy brains, spatially tuned hippocampal place cells and entorhinal grid cells exhibit distinct spike patterns in different environments, a circuit function called "remapping." We tested remapping in amyloid precursor protein knockin (APP-KI) mice with impaired spatial memory. CA1 neurons, including place cells, showed disrupted remapping, although their spatial tuning was only mildly diminished. Medial entorhinal cortex (MEC) neurons severely lost their spatial tuning and grid cells were almost absent. Fast gamma oscillatory coupling between the MEC and CA1 was also impaired. Mild disruption of MEC grid cells emerged in younger APP-KI mice, although the spatial memory and CA1 remapping of the animals remained intact. These results point to remapping impairment in the hippocampus, possibly linked to grid cell disruption, as circuit mechanisms underlying spatial memory impairment in AD.

Development of an effective alternative model for in vivo hypnozoite-induced relapse infection: A Japanese macaque (Macaca fuscata) model experimentally infected with Plasmodium cynomolgi.

In the present study, we demonstrate that the Japanese macaque (Macaca fuscata) can be used as an effective alternative in vivo model for investigating hypnozoite-induced relapsing infection caused by Plasmodium cynomolgi B strain, and that this model is comparable to the rhesus macaque model. Two female Japanese macaques (JM-1 and JM-2; aged 5 years; weighing about 4.0 kg) were used for the experiment. To produce sporozoites in mosquitoes, blood infected with P. cynomolgi B strain was collected from the donor monkey JM-1 and fed to approximately 200 mosquitoes using the standard artificial membrane feeding method. The isolated sporozoites (2 × 105) were intravenously inoculated into the JM-2 monkey, and the blood stage of the parasite was detected on day 8 after the infection. Chloroquine sulfate (CQ) was intramuscularly administered at a dosage of 6.0 mg/kg into the JM-2 monkey for 6 consecutive days from day 12 onward, after which the parasites disappeared from the peripheral blood. The first relapse occurred on day 26, which was treated again with CQ. Then, the second relapse occurred on day 44, which was cured by CQ treatment followed by the administration of primaquine phosphate (PQ) at a dosage of 1.0 mg/kg/day for 15 days. The JM-2 monkey was observed until 69 days after PQ administration, and there was no relapse during the entire follow-up period. We propose that the Japanese macaque model could contribute not only to drug screening for anti-hypnozoite activity, but could also be used as a powerful tool for investigating hypnozoite biology.