Chizuko Inui-Yamamoto

The brain mapping of the retrieval of conditioned taste aversion memory using manganese-enhanced magnetic resonance imaging in rats.

Manganese-enhanced MRI (MEMRI) is a newly developed noninvasive imaging technique of brain activities. The signal intensity of MEMRI reflects cumulative activities of the neurons. To validate the use of MEMRI technique to investigate the neural mechanisms of learning and memory, we tried to map brain areas involved in the retrieval of conditioned taste aversion (CTA) memory. CTAs were established to saccharin (conditioned stimulus: CS) by pairing its ingestion with an i.p. injection of LiCl (unconditioned stimulus: US). LiCl solutions (as a robust aversion chemical) of 0.15 M were injected i.p. 15 min after drinking the saccharine solution (CS). After the two times conditionings, these rats showed a robust aversion to the saccharine solution (CS). Rats of the control group were injected saline i.p. instead of LiCl solutions. The MRI signal intensities at the gustatory cortex (GC), the core subregion of the nucleus accumbens (NAcC), the shell subregion of the nucleus accumbens (NAcSh), the ventral pallidum (VP), the central nucleus of amygdala (CeA), the lateral hypothalamus (LH), and the basolateral nucleus of amygdala (BLA) of the conditioned group were higher than those of the control group. There were no significant differences between the conditioned and the control groups in the intensities for other regions, such as the striatum area, motor cortex, cingulate cortex, interstitial nucleus of the posterior limb of the anterior commissure and hippocampus. These indicate that the GC, NAcC, NAcSh, VP, CeA, LH and BLA have important roles in the memory retrieval of CTA.

Activation of projective neurons from the nucleus accumbens to ventral pallidum by a learned aversive taste stimulus in rats: a manganese-enhanced magnetic resonance imaging study.

Conditioned taste aversion (CTA) causes a palatability shift of a taste stimulus (conditioned stimulus, CS) from ingestive to aversive. We previously found that the ventral pallidum (VP) mediates the palatability shift in CTA. Because the VP receives major projections from the nucleus accumbens (NAc), we examined whether the presentation of CS activates the NAc-VP projective neurons after the establishment of CTA, using a manganese-enhanced magnetic resonance imaging technique. Rats were implanted with a guide cannula in the NAc and an intraoral cannula. After the surgery, they received a pairing of 5 mM saccharin solution with an i.p. injection of 0.15 M lithium chloride (CTA group) or saline (sham group). Two days after the conditioning, rats were microinjected with manganese chloride (MnCl2) into the NAc. Thirty minutes later, the rats were presented with saccharin (CTA-CS and sham-CS groups) or water (CTA-DW and sham-DW groups) via the intraoral cannula. Only the CTA-CS group showed a robust aversion to the CS. At 1 and 2 h after the MnCl2 injection, T1-weighted MR images were acquired using an 11.7 T MRI. Imaging analysis showed that significantly more manganese moved toward the VP in the CTA-CS group than in the other groups. These results indicate that the conditioned aversive taste enhanced the activities of the projective NAc-VP neurons and suggest specific involvement of the NAc-VP pathway in the rejection of CS in retrieval of CTA.

Hedonics of taste influence the gastric emptying in rats

It is known that taste sensation plays important roles in various functions including appetite, nutriental choices, food intake and digestion. To study the modulation of digestive functions by taste stimulation, we measured the gastric emptying of experimental foods with appetitive or aversive tastes in Wistar male rats. Rats were randomly divided into 3 groups: each group was trained to eat a mash mixture (8 g) made up with powdered food (4 g) and a solution (4 g) for 30 min per day. The solution was either distilled water, saccharin solution as a palatable taste, or quinine solution as an aversive taste. On the test day, the contents of the stomach of each rat were measured before and 30, 150, or 300 min after the start of eating the mash. The results showed that the food output from the stomach was increased by the palatable mash and was decreased by the aversive mash in comparison with non-adulterated mash 150 min after eating. There were no significant differences in gastric emptying among the 3 groups after deafferentation of the two peripheral taste nerve branches, the chorda tympani and glossopharyngeal nerves, or after intragastric infusions of each mash. After the establishment of conditioned taste aversions to saccharin, saccharin mash became ineffective in increasing gastric emptying. These results show that hedonically positive tastes increase, but hedonically negative tastes decrease gastric function in terms of gastric emptying.

Activation of efferents from the basolateral amygdala during the retrieval of conditioned taste aversion.

The basolateral amygdala (BLA) is critical in the retrieval of conditioned taste aversion (CTA). Although BLA neurons have axonal connections with several brain regions, it is unclear which efferent pathways are functional in CTA. The present study investigated the involvement of efferents from BLA in CTA retrieval with manganese (Mn(2+))-enhanced magnetic resonance imaging (MEMRI). Rats receiving intraoral saccharin infusion paired with intraperitoneal administration of lithium chloride (LiCl) were presented with saccharin (C-S and BC-S groups) or water (C-W group) on the test day. The BC-S group was administered with LiCl 15 min before saccharin presentation on the conditioning day (backward conditioning, BC). Another two groups were injected with saline (S-S and S-W groups) instead of LiCl. On the test day, 50 nL of 40-mM manganese chloride (MnCl2) was injected into BLA before the intraoral fluid infusion. Using MRI, we analyzed Mn(2+) movements, which indicated the activation of efferent neurons. The C-S group showed the highest activities in several efferents from BLA. Of them, the activities of the efferents to the nucleus accumbens core (NAcC), the anterior part of the bed nucleus of the stria terminalis (aBNST), and the central amygdala (CeA) were larger in the C-S group than in the Q group, which was presented with a normally aversive quinine solution. Although rats equivalently rejected conditioned aversive saccharin and quinine, the aversive responses in the C-S group, and not the Q group, were due to CTA retrieval. Therefore, our results indicated that BLA efferents to NAcC, aBNST, and CeA were specifically activated during CTA retrieval, suggesting that these efferents are key components in the neural mechanisms of CTA.

Taste preference changes throughout different life stages in male rats

Taste preference, a key component of food choice, changes with aging. However, it remains unclear how this occurs. To determine differences in taste preference between rats in different life stages, we examined the consumption of taste solutions and water using a two-bottle test. Male Sprague-Dawley rats of different ages were used: juvenile (3–6 weeks), young adult (8–11 weeks), adult (17–20 weeks), middle-aged (34–37 weeks), and old-aged (69–72 weeks). The intakes of the high and low concentration solutions presented simultaneously were measured. We observed that the old-aged group had lower preference ratios for 0.3 M sucrose and 0.1 M MSG in comparison with other groups. The preference ratio for 0.03 mM QHCl was higher in the middle-aged group than in the three younger groups and higher in the old-aged group than the juvenile group. The taste preferences for HCl and NaCl did not significantly differ among the age groups. The old-aged group tended to prefer high concentrations of sucrose, QHCl, NaCl, and MSG to low concentrations, indicating age-related decline in taste sensitivity. We also aimed to investigate differences between life stages in the electrophysiological responses of the chorda tympani nerve, one of the peripheral gustatory nerves, to taste stimuli. The electrophysiological recordings showed that aging did not alter the function of the chorda tympani nerve. This study showed that aging induced alterations in taste preference. It is likely that these alterations are a result of functional changes in other peripheral taste nerves, the gastrointestinal system, or the central nervous system.

11.7 T MR Imaging Revealed Dilatation of Virchow-Robin Spaces within Hippocampus in Maternally Lipopolysaccharide-exposed Rats.

Purpose: 11.7 Tesla MRI was examined to detect Virchow–Robin spaces (VRSs) smaller than 100 μm in the rat brain. The effects of maternal exposure to lipopolysaccharide (LPS) were evaluated on basis of the number of dilated VRSs in the offspring rat brain. Methods: T2-weighted MRI with an in-plane resolution up to 78 μm (repetition time = 5000 ms, echo time = 35 ms, slice thickness = 250 μm, imaging plane, coronal) was applied to identify VRSs. The dilated VRSs were counted in the rat brain at 5 and 10 weeks of age. The dams of half the number in each group were treated with LPS during pregnancy, and the remaining half was employed as control. LPS injection in gestation period was used to simulate maternal infections, the method of which was widely accepted as a rat model inducing neuropsychiatric disorders in the offspring. Effects of LPS exposure on the offspring rat brain were statistically investigated. Results: VRSs as small as 78 μm were successfully detected by the ultra high-field MRI. All dilated VRSs were observed within lacunosum molecular layer of hippocampus, and molecular and granular layers of dentate gyrus around hippocampal fissure. In juvenile rats (5 weeks of age), the number of dilated VRSs was significantly increased in the prenatal LPS exposed rat brain (12.9 ± 2.4, n = 7) than in the control (5.3 ± 1.5, n = 7, P < 0.05), while in young adult rats (10 weeks of age), there was no significant difference in the number between the prenatal LPS exposed rat brain (3.6 ± 0.9, n = 5) and the control (2.6 ± 0.4, n = 5). Conclusion: The results of the present study suggested that maternal infection might cause dilatation of VRSs through neural damages especially in the dentate gyrus of the offspring rats. Thus, ultra high-field MRI can offer a promising diagnostic tool capable of determining the location of neonatal brain damage caused by maternal infections.

Polymer-brush-afforded SPIO Nanoparticles Show a Unique Biodistribution and MR Imaging Contrast in Mouse Organs

Introduction: To investigate the biodistribution and retention properties of the new super paramagnetic iron oxide (new SPIO: mean hydrodynamic diameter, 100 nm) nanoparticles, which have concentrated polymer brushes in the outer shell and are difficult for phagocytes to absorb, and to compare the new SPIO with clinically approved SPIO (Resovist: mean hydrodynamic diameter, 57 nm). Materials and Methods: 16 male C57BL/6N mice were divided in two groups according to the administered SPIO (n = 8 for each group; intravenous injection does, 0.1 ml). In vivo magnetic resonance imaging (MRI) was performed before and one hour, one day, one week and four weeks after SPIO administration by two dimensional-the fast low angle shot (2D-FLASH) sequence at 11.7T. Ex vivo high-resolution images of fixed organs were also obtained by (2D-FLASH). After the ex vivo MRI, organs were sectioned and evaluated histologically to confirm the biodistribution of each particle precisely. Results: The new SPIO was taken up in small amounts by liver Kupffer cells and showed a unique in vivo MRI contrast pattern in the kidneys, where the signal intensity decreased substantially in the boundaries between cortex and outer medulla and between outer and inner medulla. We found many round dark spots in the cortex by ex vivo MRI in both groups. Resovist could be detected almost in the cortex. The shapes of the dark spots were similar to those observed in the new SPIO group. Transmission electron microscopy revealed that Resovist and the new SPIO accumulated in different cells of glomeruli, that is, endothelial and mesangial cells, respectively. Conclusion: The new SPIO was taken up in small amounts by liver tissue and showed a unique MRI contrast pattern in the kidney. The SPIO were found in the mesangial cells of renal corpuscles. Our results indicate that the new SPIO may be potentially be used as a new contrast agent for evaluation of kidney function as well as immunune function.

c-Fos expression in the parabrachial nucleus following intraoral bitter stimulation in the rat with dietary-induced zinc deficiency

Zinc deficiency causes various symptoms including taste disorders. In the present study, changes in expression of c-Fos immunoreactivity in neurons of the parabrachial nucleus (PBN), one of the relay nuclei for transmission of gustatory information, after bitter stimulation to the dorsal surface of the tongue were examined in zinc-deficient rats. Experimental zinc-deficient animals were created by feeding a low-zinc diet for 4weeks, and showed the following symptoms of zinc deficiency: low body weight, low serum zinc content and behavioral changes to avoid bitter stimulation. In normal control animals, intraoral application of 1mM quinine caused increased numbers of c-Fos-immunoreactive (c-Fos-IR) neurons in the external lateral subnucleus and external medial subnucleus of the PBN (elPBN and emPBN, respectively) compared with application of distilled water. However, in the zinc-deficient animals, the numbers of c-Fos-IR neurons in the elPBN and emPBN did not differ significantly between application of quinine and distilled water. After feeding the zinc-deficient animals a normal diet for 4weeks, the symptoms of zinc deficiency recovered, and the expression of c-Fos-IR neurons following intraoral bitter stimulation became identical to that in the normal control animals. The present results indicate that dietary zinc deficiency causes alterations to neuronal activities in the gustatory neural circuit, and that these neuronal alterations can be reversed by changing to a normal diet.

Short-Term Exposure to a Calorically Dense Diet Alters Taste-Evoked Responses in the Chorda Tympani Nerve, But Not Unconditioned Lick Responses to Sucrose

Upon presentation of a calorically dense diet, rats display hyperphagia driven by increased meal size. The increased meal size and hyperphagia are most robust across the first several days of diet exposure before changes in body weight are evident, thus it is plausible that one of the factors that drives the hyperphagia may be enhanced orosensory responsivity. Here, electrophysiological responses to an array of taste stimuli were recorded from the chorda tympani nerve, a branch of the facial nerve that innervates taste receptors in the anterior tongue, of rats presented a high-energy (45% fat and 17% sucrose) diet for 3 days. Responses in the high-energy diet group were significantly higher for 0.01, 0.03, 0.06 and 0.3 M sucrose; 0.05 M Na-saccharin; and 0.01 M quinine compared with those of chow-fed controls. Another cohort of animals was tested in 30-min brief-access taste sessions (10-s trials) to a sucrose concentration series across the first 6 days of high-energy diet presentation. Both groups responded in a concentration-dependent manner. No significant group differences in unconditioned licking or trials initiated were revealed. Results from a third cohort of rats showed that responses to sucrose in a brief-access taste test also remained largely unchanged as a function of 3-day access to a sucrose solution. Taken together, these findings suggest that 3 days of high-energy diet exposure results in alterations to peripheral gustatory signaling yet these changes do not necessarily generalize to changes in responsiveness to sucrose, as least as measured in this procedure.

Changes in regional brain temperature induced by increase in neuronal activity in rats

The brain temperature largely depends on the metabolic activities of brain tissues and blood flow in the brain. However the physiological significance of brain temperature and its regulation remain unclear. To elucidate the relationship among regional brain temperatures (rBT), blood flow and brain activity, we simultaneously recorded brain temperature, blood flow and electrophysiological neuronal activities in the gustatory cortical area (GC) of rats, in response to somatosensory stimuli (electrical stimuli, L-menthol and capsaicin) or taste stimuli to the tongue. The temperature in the GC rose by 0.01 ~ 0.05°C after the neuronal activation by electrical and taste stimuli (0.5 M NaCl, 0.5 M sucrose, or 0.1 M citric acid). The effect was stronger with L-menthol and capsaicin than that with electrical and taste stimuli. The increase of temperature in the GC was correlated with the increase of neuronal activity, but not in other area, such as the visual cortex. Furthermore, the stimulation-induced increases in spike frequency of a small neuronal ensemble in the GC were co-localized with increases in blood flow. The increases in the blood flow were showed soon before or the almost same time as the changes in the rBTs. These results suggest that the changes in the rBTs are directly related to the brain activities including the stimulation-induced neuronal activity and blood flow.