Measurement of changes in endogenous serotonin level by positron emission tomography with [18F]altanserin.

OBJECTIVE: Positron emission tomography (PET) has been used to investigate changes in the concentration of endogenous neurotransmitters. Recently, this technique has been applied to the imaging of serotonin2A receptors using [18F]altanserin. In these measurements, a reduction in binding potential (BP) suggests an increase in endogenous serotonin levels caused by pharmacological or cognitive stimulations, and the sensitivity of BP reduction depends on the characteristics of [18F]altanserin. In this study, we evaluated an analytical method for estimating the changes in endogenous serotonin levels based on PET scans with [18F]altanserin at baseline and stimulated states and validated it using simulations and small animal PET studies. METHODS: First, in the simulations, the time-activity curves at baseline and the stimulated states were generated using an extended compartment model including the competition for the receptors between the administered [18F]altanserin and endogenous serotonin. In the stimulated state, the magnitude and onset of the endogenous serotonin elevation were altered to varying degrees. In these time-activity curves, BP was estimated using the simplified reference tissue model (SRTM), and the reduction in BP was evaluated by comparison with that of the baseline state. Next, the proposed method was applied to mouse PET studies. Endogenous serotonin levels were elevated by treatment with selective serotonin reuptake inhibitors (SSRIs), and PET studies were performed twice, once with and once without treatment. In both scans, BP was estimated using the SRTM with the cerebellum as a reference region, and the reduction in BP after SSRI treatment was evaluated. RESULTS: In the simulations, the BP estimate of the stimulated state was smaller than that of the baseline state, and their reduction was related to the amount of change in the serotonin concentration. BP reduction was also affected by the onset of serotonin elevation. In the mouse studies, the BP of the cerebral cortex decreased in the scans with SSRI treatment. CONCLUSIONS: The reduction in BP estimated using the SRTM from [18F]altanserin-PET studies at baseline and in stimulated states can detect changes in the binding conditions of serotonin2A receptors. This may be useful for investigating the elevation of endogenous serotonin levels caused by stimulations.

Food Search Strategy Changes in Caenorhabditis elegans under Chronic Starvation Conditions.

Starvation is a primary threat to survival in nature. This study investigated the effects of starvation on animal behavior and neural function using a nematode model. Nematodes exhibit chemotactic responses to various compounds, including diacetyl produced by food bacteria. Locomotion, chemotactic behavior, and olfactory adaptation were measured following chronic starvation. Our results revealed a starvation-dependent reduction in locomotor activity. Chemotaxis response to the odorant diacetyl was attenuated after 2-38 hr of starvation. However, chemotactic behavior increased significantly after 48 hr of starvation compared with that after 38 hr of starvation, suggesting that food search behavior was enhanced after 48 hr of starvation. Inhibition of diacetyl adaptation was observed in the nematodes after 48 hr of starvation. However, exogenous exposure to serotonin during 48 hr of starvation caused the inhibition of diacetyl adaptation to be attenuated in following 24 hr period of normal feeding.Therefore, the inhibitory effects of starvation on olfactory adaptation may reduce chemotaxis response to the odorant diacetyl in a manner mediated by serotonin.

Inhibitory effects of caffeine on gustatory plasticity in the nematode Caenorhabditis elegans.

The effects of caffeine on salt chemotaxis learning were investigated using the nematode Caenorhabditis elegans. To estimate the degree of salt chemotaxis learning, nematodes were placed in a mixed solution of NaCl and caffeine, and then the chemotaxis index of NaCl was obtained from the nematodes placed on agar medium after pre-exposure to caffeine concentrations of 0.01, 0.1, 0.3, and 1.0%. Locomotor activity and preference behavior for caffeine were also estimated under these caffeine conditions. Nematodes pre-exposed to 0.3% caffeine showed inhibition of salt chemotaxis learning. Additional experiments indicated that nematodes showed a preference response to the middle concentration of caffeine (0.1%), with preference behavior declining in the 0.3% caffeine condition. Stable locomotor activity was observed under 0.01-0.3% caffeine conditions. These results suggest that salt chemotaxis learning with 0.3% caffeine is useful for investigating the effects of caffeine on learning in nematodes.

Long-term effects of cerebral hypoperfusion on neural density and function using misery perfusion animal model.

We investigated the chronic effects of cerebral hypoperfusion on neuronal density and functional hyperemia using our misery perfusion mouse model under unilateral common carotid artery occlusion (UCCAO). Neuronal density evaluated 28 days after UCCAO using [(11)C]flumazenil-PET and histology indicated no neurologic deficit in the hippocampus and neocortex. CBF response to sensory stimulation was assessed using laser-Doppler flowmetry. Percentage changes in CBF response of the ipsilateral hemisphere to UCCAO were 18.4 ± 3.0%, 6.9 ± 2.8%, 6.8 ± 2.3% and 4.9 ± 2.4% before, and 7, 14 and 28 days after UCCAO, respectively. Statistical significance was found at 7, 14 and 28 days after UCCAO (P < 0.01). Contrary to our previous finding (Tajima et al. 2014) showing recovered CBF response to hypercapnia on 28 days after UCCAO using the same model, functional hyperemia was sustained and became worse 28 days after UCCAO.

Vasodilation Mechanism of Cerebral Microvessels Induced by Neural Activation under High Baseline Cerebral Blood Flow Level Results from Hypercapnia in Awake Mice.

OBJECTIVE: We investigated the effects of the baseline CBF level at resting state on neurovascular coupling. METHODS: Diameters of arterioles, capillaries, and venulas in awake mouse brain were measured by a two-photon microscope. Vasodilation in each of the cerebral vessels was caused by three experimental conditions: (1) sensory stimulation, (2) 5% CO2 inhalation (hypercapnia), (3) simultaneous exposure to sensory stimulation and 5% CO2 inhalation. CBF and CBV were also measured by a microscope and a CCD camera. RESULTS: Increases in CBF and CBV were observed under all experimental conditions. After the increases in CBF and CBV due to hypercapnia, additional increases in CBF and CBV occurred during sensory stimulation. Diameter changes in arterioles were significantly larger than those in capillaries and venulas under both sensory stimulation and 5% CO2 inhalation. Additional vasodilation from sensory stimulation was observed under hypercapnia. The diameter change in each vessel type during sensory stimulation was maintained under simultaneous exposure to sensory stimulation and hypercapnia. CONCLUSIONS: The diameter change of cerebral vessels during neural activation is reproducible regardless of whether baseline CBF has increased or not. Our finding directly demonstrates the concept of uncoupling between energy consumption and energy supply during cortical activation.

Extension of the established period of diacetyl adaptation by oxygen intermediates in the nematode Caenorhabditis elegans.

After pre-exposure to the odorant diacetyl, the nematode Caenorhabditis elegans showed a decline in chemotactic responses to diacetyl, a phenomenon known as diacetyl adaptation. In the present study, we found that the established period of diacetyl adaptation in nematodes increased with the breeding temperature. When wild-type (N2) nematodes were bred at 15°C, adaptation was observed from the young adult (YA) to the 3-day-old adult that is reached 3 days after the YA stage. On breeding nematodes at 20°C and 25°C, adaptation was observed between the YA and 5-day-old adult and between the YA and the 7-day-old adult, respectively. Breeding temperature has been shown to correlate with the rate of aging in nematodes, which is related to the level of oxygen consumption. Accordingly, long-lived isp-1 and clk-1 mutants that demonstrate decreased levels of oxygen consumption showed a shorter established period of adaptation than N2 nematodes, whereas short-lived gas-1 and mev-1 mutants that have a hypersensitive response to oxygen showed a longer period of adaptation than the N2. Moreover, the established period of diacetyl adaptation in N2 nematodes was shortened by the antioxidant α-lipoic acid. These results suggest that oxygen intermediates, which are produced by oxygen consumption, play a significant role in diacetyl adaptation. Although this is only one of many factors that regulate diacetyl adaptation, such as the release of neurotransmitters and changes in intracellular conditions, the acquisition of this adaptation requires an increase in the intensity of moderate oxygen signals.

Development of new optical imaging systems of oxygen metabolism and simultaneous measurement in hemodynamic changes using awake mice.

BACKGROUND: PET allows the measurement of CBF, CBV and CMRO2 in human and plays an important role in the diagnosis of pathologic conditions and clinical research. On the other hand, in animal studies, there is no optical imaging system for evaluating changes in CBF and CBV, and oxygen metabolism, from the same brain area under awake condition. NEW METHOD: In the present study, we developed a simultaneous measurement system of LSI and IOSI, which was verified by LDF. Moreover, to evaluate oxygen metabolism, FAI was performed from the same brain area as LSI and IOSI measurements. RESULTS: The change in CBF according to LSI was correlated with that by LDF. Similarly, the change in CBV obtained by IOSI was also correlated with RBC concentration change measured by LDF. The change in oxygen metabolism by FAI was associated with that in CBF obtained by LSI, although the change in CBF was greater than that in oxygen metabolism. COMPARISON WITH EXISTING METHOD(S): We revealed that the relationship between oxygen metabolism and CBF as measured by our system was in good agreement with the relationship between CMRO2 and CBF in human PET studies. CONCLUSIONS: Our measurement system of CBF, CBV and oxygen metabolism is not only useful for studying neurovascular coupling, but also easily corroborates human PET studies.

Cerebral hemodynamic response to acute hyperoxia in awake mice.

Cerebral hemodynamic response to acute hyperoxia was investigated in awake mice. Using laser-Doppler flowmetry (LDF), baseline cerebral blood flow (CBF) and the cerebrovascular responses to whisker stimulation were measured in awake mice during normoxia and hyperoxia. Using two-photon laser scanning microscopy (TPLSM), the changes in cortical microvasculature were measured during normoxia and hyperoxia. During hyperoxia (PaO2=482.3±19.7mmHg), baseline CBF was 6.8% lower than normoxia (PaO2=97.3±6.0mmHg). The degree of increase in CBF evoked by whisker stimulation was greater during hyperoxia (18.1±5.0%) than normoxia (13.1±3.5%) (P<0.05). TPLSM imaging of the somatosensory cortex showed vasconstriction in arterioles and capillaries during hyperoxia. Since the effective diffusivity for oxygen in the capillary bed might decrease by hyperoxia due to a decrease in capillary blood volume according to Hyder׳s model, an increase in the cerebral metabolic rate of oxygen utilization by neural activation during hyperoxia might need a greater increase in CBF as compared with normoxia. The hemodynamic response to neural activation could be modified by acute hyperoxia due to modification of the relation between changes in CBF and oxygen consumption by neural activation.

Inhibition of gustatory plasticity due to acute nicotine exposure in the nematode Caenorhabditis elegans.

The present study investigated the effect of nicotine exposure on gustatory plasticity in the nematode Caenorhabditis elegans. The chemotactic response of wild-type N2 nematodes pre-exposed to 100mM NaCl with 3.0mM nicotine was almost the same as that of mock-conditioned nematodes unexposed to NaCl; however, the response of N2 nematodes pre-exposed to NaCl without nicotine was significantly lower than that of mock-conditioned nematodes. These results indicate that gustatory plasticity is inhibited by acute nicotine exposure. Inhibition of gustatory plasticity was observed when cat-2 mutants with a defect in dopamine biosynthesis were pre-exposed to NaCl with 3.0mM nicotine. Acute nicotine exposure did not cause inhibition of gustatory plasticity in bas-1 mutants, which had defects in both serotonin and dopamine secretion, and tph-1 mutants, which had a defect in serotonin secretion only. However, inhibition of gustatory plasticity was observed when bas-1 and tph-1 mutants were maintained on a growth plate that included serotonin. These results suggest that serotonin signaling plays an essential role in the modulation of the acute effects of nicotine.

The role of oxygen intermediates in the retention time of diacetyl adaptation in the nematode Caenorhabditis elegans.

Continuous presentation of the odorant diacetyl to the nematode Caenorhabditis elegans causes a decrease in the level of chemotactic response to diacetyl. This decline in response is caused by diacetyl adaptation. When wild-type nematodes were maintained at 15°C after pre-exposure to diacetyl, diacetyl adaptation did not continue up to 2 hr. Adaptation continued up to 6 hr in nematodes bred at 20°C, and it continued beyond 12 hr in nematodes bred at 25°C. These results indicate that the retention time of diacetyl adaptation is dependent on the environmental breeding temperature and suggest that moderate oxygen signals are required for maintaining the attenuated response to diacetyl because of the correlation between breeding temperature and production of oxygen intermediates. When isp-1 and clk-1 mutants, which show reduced rates of oxygen intermediate production, were maintained at 20 and 25°C after pre-exposure to diacetyl, the mutants showed a shorter retention time of diacetyl adaptation compared with that of wild-type nematodes. When gas-1 and mev-1 mutants, which have a hypersensitive response to oxidative stress, were maintained at 15 and 20°C, they showed a longer retention time of adaptation, that is, adaptation continued beyond 2 and 12 hr, respectively. When wild-type nematodes were maintained on plates that included 0.05% α-lipoic acid, which suppresses production of oxygen intermediates, the retention time of adaptation did not continue up to 6 hr in nematodes bred at 20°C and up to 12 hr in nematodes bred at 25°C. These results support the possibility that oxygen intermediates contribute to retention time for diacetyl adaptation in the nematode C. elegans.