Odor-evoked layer-specific fMRI activities in the awake mouse olfactory bulb.

Awake rodent fMRI is increasingly common over the use of anesthesia since it permits behavioral paradigms and does not confound normal brain function or neurovascular coupling. It is well established that adequate acclimation to the loud fMRI environment and head fixation reduces stress in the rodents and allows for whole brain imaging with little contamination from motion. However, it is unknown whether high-resolution fMRI with increased susceptibility to motion and lower sensitivity can measure small, but spatially discrete, activations in awake mice. To examine this, we used contrast-enhanced cerebral blood volume-weighted (CBVw) fMRI in the mouse olfactory bulb for its enhanced sensitivity and neural specificity. We determined that activation patterns in the glomerular layer to four different odors were spatially distinct and were consistent with previously established histological patterns. In addition, odor-evoked laminar activations were greatest in superficial layers that decreased with laminar depth, similar to previous observations. Interestingly, the fMRI response strengths in the granule cell layer were greater in awake mice than our previous anesthetized rat studies, suggesting that feedback neural activities were intact with wakefulness. We finally determined that fMRI signal changes to repeated odor exposure (i.e., olfactory adaptation) attenuated relatively more in the feedback granule cell layer compared to the input glomerular layer, which is consistent with prior observations. We, therefore, conclude that high-resolution CBVw fMRI can measure odor-specific activation patterns and distinguish changes in laminar activity of head and body restrained awake mice.

Heat Stroke Prevention in Hot Specific Occupational Environment Enhanced by Supervised Machine Learning with Personalized Vital Signs.

Recently, wet-bulb globe temperature (WBGT) has attracted a lot of attention as a useful index for measuring heat strokes even when core body temperature cannot be available for the prevention. However, because the WBGT is only valid in the vicinity of the WBGT meter, the actual ambient heat could be different even in the same room owing to ventilation, clothes, and body size, especially in hot specific occupational environments. To realize reliable heat stroke prevention in hot working places, we proposed a new personalized vital sign index, which is combined with several types of vital data, including the personalized heat strain temperature (pHST) index based on the temperature/humidity measurement to adjust the WBGT at the individual level. In this study, a wearable device was equipped with the proposed pHST meter, a heart rate monitor, and an accelerometer. Additionally, supervised machine learning based on the proposed personalized vital index was introduced to improve the prevention accuracy. Our developed system with the proposed vital sign index achieved a prevention accuracy of 85.2% in a hot occupational experiment in the summer season, where the true positive rate and true negative rate were 96.3% and 83.7%, respectively.

Postsynaptic activity of inhibitory neurons evokes hemodynamic fMRI responses.

Functional MRI responses are localized to the synaptic sites of evoked inhibitory neurons, but it is unknown whether, or by what mechanisms, these neurons initiate functional hyperemia. Here, the neuronal origins of these hemodynamic responses were investigated by fMRI or local field potential and blood flow measurements during topical application of pharmacological agents when GABAergic granule cells in the rat olfactory bulb were synaptically targeted. First, to examine if postsynaptic activation of these inhibitory neurons was required for neurovascular coupling, we applied an NMDA receptor antagonist during cerebral blood volume-weighted fMRI acquisition and found that responses below the drug application site (up to ~1.5 mm) significantly decreased within ~30 min. Similarly, large decreases in granule cell postsynaptic activities and blood flow responses were observed when AMPA or NMDA receptor antagonists were applied. Second, inhibition of nitric oxide synthase preferentially decreased the initial, fast component of the blood flow response, while inhibitors of astrocyte-specific glutamate transporters and vasoactive intestinal peptide receptors did not decrease blood flow responses. Third, inhibition of GABA release with a presynaptic GABAB receptor agonist caused less reduction of neuronal and blood flow responses compared to the postsynaptic glutamate receptor antagonists. In conclusion, local hyperemia by synaptically-evoked inhibitory neurons was primarily driven by their postsynaptic activities, possibly through NMDA receptor-dependent calcium signaling that was not wholly dependent on nitric oxide.

α-Emitting cancer therapy using 211 At-AAMT targeting LAT1.

α-Methyl-l-tyrosine (AMT) has a high affinity for the cancer-specific l-type amino acid transporter 1 (LAT1). Therefore, we established an anti-cancer therapy, with 211 At-labeled α-methyl-l-tyrosine (211 At-AAMT) as a carrier of 211 At into tumors. 211 At-AAMT had high affinity for LAT1, inhibited tumor cell growth, and induced DNA double-stranded breaks in vitro. We evaluated the accumulation of 211 At-AAMT in vivo and the role of LAT1. Treatment with 0.4 MBq/mouse 211 At-AAMT inhibited tumor growth in the PANC-1 tumor model and 1 MBq/mouse 211 At-AAMT inhibited metastasis in the lung of the B16F10 metastasis model. Our results suggested that 211 At would be useful for anti-cancer therapy and that LAT1 is suitable as a target for radionuclide therapy.

Chemical exchange sensitive MRI of glucose uptake using xylose as a contrast agent.

PURPOSE: Glucose and its analogs can be detected by CEST and chemical exchange spin-lock (CESL) MRI techniques, but sensitivity is still a bottleneck for human applications. Here, CESL and CEST sensitivity and the effect of injection on baseline physiology were evaluated for a glucose analog, xylose. METHODS: The CEST and CESL sensitivity were evaluated at 9.4 T in phantoms and by in vivo rat experiments with 0.5 and 1 g/kg xylose injections. Arterial blood glucose level was sampled before and after 1 g/kg xylose injection. The effect of injection on baseline neuronal activity was measured by electrophysiology data during injections of saline, xylose, and 2-deoxy-D-glucose. RESULTS: In phantoms, xylose shows similar chemical exchange sensitivity and pH-dependence with that of glucose. In rat experiments with a bolus injection, CESL shows higher sensitivity in the detection of xylose than CEST, and the sensitivity of xylose is much higher than glucose. Injection of xylose does not significantly affect blood glucose level and baseline neural activity for 1-g/kg and 0.6-g/kg doses, respectively. CONCLUSION: Due to its relatively high sensitivity and safety, xylose is a promising contrast agent for the study of glucose uptake.

Foundations of layer-specific fMRI and investigations of neurophysiological activity in the laminarized neocortex and olfactory bulb of animal models.

Laminar organization of neuronal circuits is a recurring feature of how the brain processes information. For instance, different layers compartmentalize different cell types, synaptic activities, and have unique intrinsic and extrinsic connections that serve as units for specialized signal processing. Functional MRI is an invaluable tool to investigate laminar processing in the in vivo human brain, but it measures neuronal activity indirectly by way of the hemodynamic response. Therefore, the accuracy of high-resolution laminar fMRI depends on how precisely it can measure localized microvascular changes nearest to the site of evoked activity. To determine the specificity of fMRI responses to the true neurophysiological responses across layers, the flexibility to invasive procedures in animal models has been necessary. In this review, we will examine different fMRI contrasts and their appropriate uses for layer-specific fMRI, and how localized laminar processing was examined in the neocortex and olfactory bulb. Through collective efforts, it was determined that microvessels, including capillaries, are regulated within single layers and that several endogenous and contrast-enhanced fMRI contrast mechanisms can separate these neural-specific vascular changes from the nonspecific, especially cerebral blood volume-weighted fMRI with intravenous contrast agent injection. We will also propose some open questions that are relevant for the successful implementation of layer-specific fMRI and its potential future directions to study laminar processing when combined with optogenetics.

Dominance of layer-specific microvessel dilation in contrast-enhanced high-resolution fMRI: Comparison between hemodynamic spread and vascular architecture with CLARITY.

Contrast-enhanced cerebral blood volume-weighted (CBVw) fMRI response peaks are specific to the layer of evoked synaptic activity (Poplawsky et al., 2015), but the spatial resolution limit of CBVw fMRI is unknown. In this study, we measured the laminar spread of the CBVw fMRI evoked response in the external plexiform layer (EPL, 265 ± 65 µm anatomical thickness, mean ± SD, n = 30 locations from 5 rats) of the rat olfactory bulb during electrical stimulation of the lateral olfactory tract and examined its potential vascular source. First, we obtained the evoked CBVw fMRI responses with a 55 × 55 µm2 in-plane resolution and a 500-µm thickness at 9.4 T, and found that the fMRI signal peaked predominantly in the inner half of EPL (136 ± 54 µm anatomical thickness). The mean full-width at half-maximum of these fMRI peaks was 347 ± 102 µm and the functional spread was approximately 100 or 200 µm when the effects of the laminar thicknesses of EPL or inner EPL were removed, respectively. Second, we visualized the vascular architecture of EPL from a different rat using a Clear Lipid-exchanged Anatomically Rigid Imaging/immunostaining-compatible Tissue hYdrogel (CLARITY)-based tissue preparation method and confocal microscopy. Microvascular segments with an outer diameter of <11 µm accounted for 64.3% of the total vascular volume within EPL and had a mean segment length of 55 ± 40 µm (n = 472). Additionally, vessels that crossed the EPL border had a mean segment length outside of EPL equal to 73 ± 61 µm (n = 28), which is comparable to half of the functional spread (50-100 µm). Therefore, we conclude that dilation of these microvessels, including capillaries, likely dominate the CBVw fMRI response and that the biological limit of the fMRI spatial resolution is approximately the average length of 1-2 microvessel segments, which may be sufficient for examining sublaminar circuits.

Inhibitory Neuron Activity Contributions to Hemodynamic Responses and Metabolic Load Examined Using an Inhibitory Optogenetic Mouse Model.

Hemodynamic signals are routinely used to noninvasively assess brain function in humans and animals. This work examined the contribution of inhibitory neuron activity on hemodynamic responses captured by changes in blood flow, volume and oxygenation in the cortex of lightly anesthetized mice. Because cortical activity is not commonly initiated by inhibitory neurons, experiments were conducted to examine the neuronal activity properties elicited by photo-stimulation. We observed comparable increases in neuronal activity evoked by forelimb and photo-stimulation; however, significantly larger increases in blood flow and volume were produced by photo-stimulation of inhibitory neurons compared with forelimb stimulation. Following blockade of glutamate and GABA-A receptors to reduce postsynaptic activity contributions, neuronal activity was reliably modulated and hemodynamic changes persisted, though slightly reduced. More importantly, photo-stimulation-evoked changes in blood flow and volume were suppressed by 75-80% with the administration of a nitric oxide synthase inhibitor, suggesting that inhibitory neurons regulate blood flow mostly via nitric oxide. Lastly, forelimb and photo-stimulation of excitatory neurons produced local decreases in blood oxygenation, while large increases were generated by photo-stimulation of inhibitory neurons. Estimates of oxygen metabolism suggest that inhibitory neuron activity has a small impact on tissue metabolic load, indicating a mismatch between the metabolic demand and blood flow regulation properties of inhibitory and excitatory neurons.

Layer-Specific fMRI Responses to Excitatory and Inhibitory Neuronal Activities in the Olfactory Bulb.

High-resolution functional magnetic resonance imaging (fMRI) detects localized neuronal activity via the hemodynamic response, but it is unclear whether it accurately identifies neuronal activity specific to individual layers. To address this issue, we preferentially evoked neuronal activity in superficial, middle, and deep layers of the rat olfactory bulb: the glomerular layer by odor (5% amyl acetate), the external plexiform layer by electrical stimulation of the lateral olfactory tract (LOT), and the granule cell layer by electrical stimulation of the anterior commissure (AC), respectively. Electrophysiology, laser-Doppler flowmetry of cerebral blood flow (CBF), and blood oxygenation level-dependent (BOLD) and cerebral blood volume-weighted (CBV) fMRI at 9.4 T were performed independently. We found that excitation of inhibitory granule cells by stimulating LOT and AC decreased the spontaneous multi-unit activities of excitatory mitral cells and subsequently increased CBF, CBV, and BOLD signals. Odor stimulation also increased the hemodynamic responses. Furthermore, the greatest CBV fMRI responses were discretely separated into the same layers as the evoked neuronal activities for all three stimuli, whereas BOLD was poorly localized with some exception to the poststimulus undershoot. In addition, the temporal dynamics of the fMRI responses varied depending on the stimulation pathway, even within the same layer. These results indicate that the vasculature is regulated within individual layers and CBV fMRI has a higher fidelity to the evoked neuronal activity compared with BOLD. Our findings are significant for understanding the neuronal origin and spatial specificity of hemodynamic responses, especially for the interpretation of laminar-resolution fMRI. SIGNIFICANCE STATEMENT: Functional magnetic resonance imaging (fMRI) is a noninvasive, in vivo technique widely used to map function of the entire brain, including deep structures, in animals and humans. However, it measures neuronal activity indirectly by way of the vascular response. It is currently unclear how finely the hemodynamic response is regulated within single cortical layers and whether increased inhibitory neuronal activities affect fMRI signal changes. Both laminar specificity and the neural origins of fMRI are important to interpret functional maps properly, which we investigated by activating discrete rat olfactory bulb circuits.

Improved spatial accuracy of functional maps in the rat olfactory bulb using supervised machine learning approach.

Functional MRI (fMRI) is a popular and important tool for noninvasive mapping of neural activity. As fMRI measures the hemodynamic response, the resulting activation maps do not perfectly reflect the underlying neural activity. The purpose of this work was to design a data-driven model to improve the spatial accuracy of fMRI maps in the rat olfactory bulb. This system is an ideal choice for this investigation since the bulb circuit is well characterized, allowing for an accurate definition of activity patterns in order to train the model. We generated models for both cerebral blood volume weighted (CBVw) and blood oxygen level dependent (BOLD) fMRI data. The results indicate that the spatial accuracy of the activation maps is either significantly improved or at worst not significantly different when using the learned models compared to a conventional general linear model approach, particularly for BOLD images and activity patterns involving deep layers of the bulb. Furthermore, the activation maps computed by CBVw and BOLD data show increased agreement when using the learned models, lending more confidence to their accuracy. The models presented here could have an immediate impact on studies of the olfactory bulb, but perhaps more importantly, demonstrate the potential for similar flexible, data-driven models to improve the quality of activation maps calculated using fMRI data.

Neural and hemodynamic responses elicited by forelimb- and photo-stimulation in channelrhodopsin-2 mice: insights into the hemodynamic point spread function.

Hemodynamic responses are commonly used to map brain activity; however, their spatial limits have remained unclear because of the lack of a well-defined and malleable spatial stimulus. To examine the properties of neural activity and hemodynamic responses, multiunit activity, local field potential, cerebral blood volume (CBV)-sensitive optical imaging, and laser Doppler flowmetry were measured from the somatosensory cortex of transgenic mice expressing Channelrhodopsin-2 in cortex Layer 5 pyramidal neurons. The magnitude and extent of neural and hemodynamic responses were modulated using different photo-stimulation parameters and compared with those induced by somatosensory stimulation. Photo-stimulation-evoked spiking activity across cortical layers was similar to forelimb stimulation, although their activity originated in different layers. Hemodynamic responses induced by forelimb- and photo-stimulation were similar in magnitude and shape, although the former were slightly larger in amplitude and wider in extent. Altogether, the neurovascular relationship differed between these 2 stimulation pathways, but photo-stimulation-evoked changes in neural and hemodynamic activities were linearly correlated. Hemodynamic point spread functions were estimated from the photo-stimulation data and its full-width at half-maximum ranged between 103 and 175 µm. Therefore, submillimeter functional structures separated by a few hundred micrometers may be resolved using hemodynamic methods, such as optical imaging and functional magnetic resonance imaging.

Feasibility studies towards future self-sufficient supply of the (99)Mo-(99m)Tc isotopes with Japanese accelerators.

In order to establish a self-sufficient supply of (99m)Tc, we studied feasibilities to produce its parent nucleus, (99)Mo, using Japanese accelerators. The daughter nucleus, (99m)Tc, is indispensable for medical diagnosis. (99)Mo has so far been imported from abroad, which is separated from fission products generated in nuclear reactors using enriched (235)U fuel. We investigated (99m)Tc production possibilities based on the following three scenarios: (1) (99)Mo production by the (n, 2n) reaction by spallation neutrons at the J-PARC injector, LINAC; (2) (99)Mo production by the (p, pn) reaction at Ep = 50-80 MeV proton at the RCNP cyclotron; (3) (99m)Tc direct production with a 20 MeV proton beam from the PET cyclotron. Among these three scenarios, scenario (1) is for a scheme on a global scale, scenario (2) works in a local area, and both cases take a long time for negotiations. Scenario (3) is attractive because we can use nearly 50 PET cyclotrons in Japan for (99m)Tc production. We here consider both the advantages and disadvantages among the three scenarios by taking account of the Japanese accelerator situation.

Parvalbumin interneuron activity induces slow cerebrovascular fluctuations in awake mice.

Neuronal regulation of cerebrovasculature underlies brain imaging techniques reliant on cerebral blood flow (CBF) changes. However, interpreting these signals requires understanding their neural correlates. Parvalbumin (PV) interneurons are crucial in network activity, but their impact on CBF is not fully understood. Optogenetic studies show that stimulating cortical PV interneurons induces diverse CBF responses, including rapid increases, decreases, and slower delayed increases. To clarify this relationship, we measured hemodynamic and neural responses to optogenetic stimulation of PV interneurons expressing Channelrhodopsin-2 during evoked and ongoing resting-state activity in the somatosensory cortex of awake mice. Two-photon microscopy (2P) Ca2+ imaging showed robust activation of PV-positive (PV+) cells and inhibition of PV-negative (PV-) cells. Prolonged PV+ cell stimulation led to a delayed, slow CBF increase, resembling a secondary peak in the CBF response to whisker stimulation. 2P vessel diameter measurements revealed that PV+ cell stimulation induced rapid arterial vasodilation in superficial layers and delayed vasodilation in deeper layers. Ongoing activity recordings indicated that both PV+ and PV- cell populations modulate arterial fluctuations at rest, with PV+ cells having a greater impact. These findings show that PV interneurons generate a complex depth-dependent vascular response, dominated by slow vascular changes in deeper layers.

Long-range inhibitory neurons mediate cortical neurovascular coupling.

To meet the high energy demands of brain function, cerebral blood flow (CBF) parallels changes in neuronal activity by a mechanism known as neurovascular coupling (NVC). However, which neurons play a role in mediating NVC is not well understood. Here, we identify in mice and humans a specific population of cortical GABAergic neurons that co-express neuronal nitric oxide synthase and tachykinin receptor 1 (Tacr1). Through whole-tissue clearing, we demonstrate that Tacr1 neurons extend local and long-range projections across functionally connected cortical areas. We show that whisker stimulation elicited Tacr1 neuron activity in the barrel cortex through feedforward excitatory pathways. Additionally, through optogenetic experiments, we demonstrate that Tacr1 neurons are instrumental in mediating CBF through the relaxation of mural cells in a similar fashion to whisker stimulation. Finally, by electron microscopy, we observe that Tacr1 processes contact astrocytic endfeet. These findings suggest that Tacr1 neurons integrate cortical activity to mediate NVC.

Spatiotemporal characteristics and vascular sources of neural-specific and -nonspecific fMRI signals at submillimeter columnar resolution.

The neural specificity of hemodynamic-based functional magnetic resonance imaging (fMRI) signals is dependent on both the vascular regulation and the sensitivity of the applied fMRI technique to different types and sizes of blood vessels. In order to examine the specificity of MRI-detectable hemodynamic responses, submillimeter blood oxygenation level-dependent (BOLD) and cerebral blood volume (CBV) fMRI studies were performed in a well-established cat orientation column model at 9.4 T. Neural-nonspecific and -specific signals were separated by comparing the fMRI responses of orthogonal orientation stimuli. The BOLD response was dominantly neural-nonspecific, mostly originating from pial and intracortical emerging veins, and thus was highly correlated with baseline blood volume. Uneven baseline CBV may displace or distort small functional domains in high-resolution BOLD maps. The CBV response in the parenchyma exhibited dual spatiotemporal characteristics, a fast and early neural-nonspecific response (with 4.3-s time constant) and a slightly slower and delayed neural-specific response (with 9.4-s time constant). The nonspecific CBV signal originates from early-responding arteries and arterioles, while the specific CBV response, which is not correlated with baseline blood volume, arises from late-responding microvessels including small pre-capillary arterioles and capillaries. Our data indicate that although the neural specificity of CBV fMRI signals is dependent on stimulation duration, high-resolution functional maps can be obtained from steady-state CBV studies.

Effects of the α2-adrenergic receptor agonist dexmedetomidine on neural, vascular and BOLD fMRI responses in the somatosensory cortex.

This article describes the effects of dexmedetomidine (DEX) - the active ingredient of medetomidine, which is the latest popular sedative for functional magnetic resonance imaging (fMRI) in rodents - on multiple unit activity, local field potential (LFP), cerebral blood flow (CBF), pial vessel diameter [indicative of cerebral blood volume (CBV)], and blood oxygenation level-dependent (BOLD) fMRI. These measurements were obtained from the rat somatosensory cortex during 10 s of forepaw stimulation. We found that the continuous intravascular systemic infusion of DEX (50 µg/kg/h, doses typically used in fMRI studies) caused epileptic activities, and that supplemental isoflurane (ISO) administration of ~0.3% helped to suppress the development of epileptic activities and maintained robust neuronal and hemodynamic responses for up to 3 h. Supplemental administration of N(2)O in addition to DEX nearly abolished hemodynamic responses even if neuronal activity remained. Under DEX + ISO anesthesia, spike firing rate and the delta power of LFP increased, whereas beta and gamma power decreased, as compared with ISO-only anesthesia. DEX administration caused pial arteries and veins to constrict nearly equally, resulting in decreases in baseline CBF and CBV. Evoked LFP and CBF responses to forepaw stimulation were largest at a frequency of 8-10 Hz, and a non-linear relationship was observed. Similarly, BOLD fMRI responses measured at 9.4 T were largest at a frequency of 10 Hz. Both pial arteries and veins dilated rapidly (artery, 32.2%; vein, 5.8%), and venous diameter returned to baseline slower than arterial diameter. These results will be useful for designing, conducting and interpreting fMRI experiments under DEX sedation.

BOLD responses to different temporal frequency stimuli in the lateral geniculate nucleus and visual cortex: insights into the neural basis of fMRI.

The neural basis of the blood oxygenation level dependent (BOLD) functional magnetic resonance imaging (fMRI) remains largely unknown after decades of research. To investigate this issue, the unique property of the temporal frequency tuning that could separate neural input and output in the primary visual cortex was used as a model. During moving grating stimuli of 1, 2, 10 and 20Hz temporal frequencies, we measured 9.4-T BOLD fMRI responses simultaneously in the primary visual cortex of area 17 (A17) and area 18 (A18), and the lateral geniculate nucleus (LGN) of isoflurane-anesthetized cat. Our results showed that preferred temporal frequencies of the BOLD responses for A17, A18 and LGN were 3.1Hz, 4.5Hz and 6.0Hz, respectively, which were comparable to the previously reported electrophysiological data. Additionally, the difference of BOLD response onset time between LGN and A17 was 0.5s, which is 18 times larger than the difference of neural activity onset time between these areas. We then compared the frequency-dependent BOLD fMRI response of A17 with tissue partial pressure of oxygen (pO(2)) and electrophysiological data of the same animal model reported by Viswanathan and Freeman (Nature Neuroscience, 2007). The BOLD tuning curve resembled the low frequency band (<12Hz) of local field potential (LFP) tuning curve rather than spiking activity, gamma band (25-90Hz) of LFP, and tissue pO(2) tuning curves, suggesting that the BOLD fMRI signal relates closer to low frequency LFP.

Time-dependent spatial specificity of high-resolution fMRI: insights into mesoscopic neurovascular coupling.

High-resolution functional magnetic resonance imaging (fMRI) is becoming increasingly popular because of the growing availability of ultra-high magnetic fields which are capable of improving sensitivity and spatial resolution. However, it is debatable whether increased spatial resolutions for haemodynamic-based techniques, like fMRI, can accurately detect the true location of neuronal activity. We have addressed this issue in functional columns and layers of animals with haemoglobin-based optical imaging and different fMRI contrasts, such as blood oxygenation level-dependent, cerebral blood flow and cerebral blood volume fMRI. In this review, we describe empirical evidence primarily from our own studies on how well these fMRI signals are spatially specific to the neuronally active site and discuss insights into neurovascular coupling at the mesoscale. This article is part of the theme issue 'Key relationships between non-invasive functional neuroimaging and the underlying neuronal activity'.

Frequency-dependent neural activity, CBF, and BOLD fMRI to somatosensory stimuli in isoflurane-anesthetized rats.

Inhalation anesthetics (e.g. isoflurane) are preferable for longitudinal fMRI experiments in the same animals. We previously implemented isoflurane anesthesia for rodent forepaw stimulation studies, and optimized the stimulus parameters with short stimuli (1-3-s long stimulation with ten electric pulses). These parameters, however, may not be applicable for long periods of stimulation because repetitive stimuli induce neural adaptation. Here we evaluated frequency-dependent responses (pulse width of 1.0 ms and current of 1.5 mA) for 30-s long stimulation under 1.3-1.5% isoflurane anesthesia. The cerebral blood flow (CBF) response (using laser Doppler flowmetry: CBF(LDF)) and field potential (FP) changes were simultaneously measured for nine stimulus frequencies (1-24 Hz). CBF (using arterial spin labeling: CBF(ASL)) and blood oxygenation level dependent (BOLD) fMRI responses were measured at 9.4 T for four stimulus frequencies (1.5-12 Hz). Higher stimulus frequencies (12-24 Hz) produced a larger FP per unit time initially, but decreased more rapidly later due to neural adaptation effects. On the other hand, lower stimulus frequencies (1-3 Hz) induced smaller, but sustained FP activities over the entire stimulus period. Similar frequency-dependencies were observed in CBF(LDF), CBF(ASL) and BOLD responses. A linear relationship between FP and CBF(LDF) was observed for all stimulus frequencies. Stimulation frequency for the maximal cumulative neural and hemodynamic changes is dependent on stimulus duration; 8-12 Hz for short stimulus durations (<10s) and 6-8 Hz for 30-s stimulation. Our findings suggest that neural adaptation should be considered in determining the somatosensory stimulation frequency and duration under isoflurane anesthesia.

Development of an MeV proton generator using a 3He ion source at the Research Center for Nuclear Physics.

For simple applications, such as the calibration of a charged particle detector, a multi-MeV proton generator may be preferable to cyclotrons or electrostatic accelerators such as Van de Graaff generator. Thus, a proton generating system, consisting of an 18 GHz superconducting (SC)-ECR ion source and a deuterated polyethylene target, was developed at the Research Center for Nuclear Physics at Osaka University. A 3He2+ beam of 400 eµA was generated by the SC-ECR ion source with the acceleration voltage of 20 kV in an experiment that utilized the fusion reaction 3He + deuteron (D) → proton(P) + 4He. Protons with energies of the order of several MeV were successfully generated at 3.67 Hz at the atmosphere side of the target in the experimental setup, using a novel target base with a thin aluminum window.