EFFECTS OF ISOFLURANE AND ALPHA-CHLORALOSE ANESTHESIA ON BOLD fMRI RESPONSES TO INGESTED L-GLUTAMATE IN RATS T. TSURUGIZAWA, A. UEMATSU, H. UNEYAMA AND K. TORII* Institute of Life Sciences, Ajinomoto, Co., Inc., Suzuki-cho 1-1. Ka- wasaki-ku, Kawasaki 210 – 8681, Japan Abstract—It is important to investigate the effect of anesthe- sia on blood oxygenation level-dependent (BOLD) signals in an animal model. Many researchers have investigated the BOLD response to visual, sensory, and chemical stimuli in anesthetized rats. There are no reports, however, comparing the differences in the BOLD signal change between anesthe- tized and conscious rats when a visceral nutrient signal arises. Here, using functional magnetic resonance imaging (fMRI), we investigated the differences in the BOLD signal changes after intragastric administration of L-glutamate (Glu) under three anesthesia conditions: conscious, alpha-chlora- lose-anesthetized, and isoflurane-anesthetized condition. Under the conscious and alpha-chloralose condition, we ob- served the significant BOLD signal increase in the medial prefrontal cortex (mPFC), insular cortex (IC), hippocampus, and several hypothalamic regions including the lateral and ventromedial nucleus. In chloralose group, however, gut Glu stimulation induced BOLD signal increase in the prelim- bic cortex and orbital cortex, which did not activate in con- scious condition. Meanwhile, under isoflurane-anesthetized condition, we did not observe the BOLD signal increase in these areas. BOLD signal intensity in the nucleus of the solitary tract (NTS), to which vagus nerve transmits the vis- ceral information from the gastrointestinal tract, increased in all conditions. Importantly, under conscious condition, we observed increased BOLD signal intensity in several regions related to the metabolic state (i.e. hunger or satiety), such as the mPFC, ventromedial and lateral hypothalamus (LH). Our results suggest that alpha-chloralose and isoflurane anesthe- sia caused distinct effects on BOLD response to the gut L-Glu stimulation in several brain regions. © 2010 IBRO. Published by Elsevier Ltd. All rights reserved. Key words: functional MRI, rat, awake, anesthesia, glutamate, nutrition. Functional magnetic resonance imaging (fMRI) is a pow- erful tool for investigating brain response in primates and rodents. Many fMRI studies in rodents have been per- formed under anesthetized condition to reduce motion ar- tifacts and restraint stress; however, there are some prob- lems with using anesthesia in fMRI measurement because it alters cell metabolism, neural activity, and cerebral blood flow (CBF) (Sicard et al., 2003; Kreuter et al., 2004). Thus, it is important to investigate the effects of anesthesia in fMRI studies. To date, some studies have revealed the effects of anesthesia on the blood oxygenation level-de- pendent (BOLD) signal response to chemical or sensory stimulation (Lahti et al., 1999; Brevard et al., 2003; Sicard et al., 2003; Luo et al., 2007). However, there are still no reports addressing the effects of anesthesia on the BOLD response related to the gut nutrient sensing in the brain. L-glutamate (Glu) is an amino acid that is an integral part of most foods and a key molecule in the cell metabolism of the gut (Lindemann, 2000). In addition to flavor enhancement in cuisine, Glu also causes a post-ingestive positive effect in humans and rats, suggesting that ingested Glu has an im- portant role in the evaluation of foods (Prescott, 2004; Ue- matsu et al., 2009). Recent studies have also revealed the existence of taste-sensing systems for Glu (e.g. T1R3 and alpha-gustducin) in the gut (Hofer et al., 1999). Intragastri- cally infused Glu evokes neural signals to the brain via the vagus nerve (Uneyama et al., 2006). While, Glu absorbed in the gut barely passes into the brain through the blood– brain barrier. These observations suggest that the vagus nerve is the main pathway to convey Glu information from the gut to the brain. Thus, Glu solutions are useful for clarifying the mechanisms of gut chemo-sensation via vagus nerve. Our previous fMRI study in alpha-chloralose anesthe- tized rats has directly shown that intragastric administra- tion of Glu elicits the BOLD signal increase in the forebrain regions including the insular cortex (IC) and hypothalamus (Tsurugizawa et al., 2008), but it is still unknown whether the BOLD response is similar to that under the other an- esthetics and conscious condition. In this study, we fo- cused on two distinct types of anesthetics: isoflurane and alpha-chloralose. These anesthetics are often used in ro- dent fMRI studies to investigate the brain regions that respond to sensory, visual, and olfactory stimuli (Blaizot et al., 2000; Chang and Shyu, 2001), although they have different effects on the BOLD response. Recent rodent studies have shown differences in CBF and the BOLD signals between anesthetized and unanesthetized rats (Nakao et al., 2001; Duong, 2007; Luo et al., 2007). These differences are derived from the effects of anesthesia on neural activity and cerebral blood vessels. Isoflurane is a volatile anesthetic that depresses neural activity and cere- bral metabolism (Hansen et al., 1989), in addition to mod- ulating dopamine release in the striatum (Adachi et al., 2005). Other reports have shown that isoflurane inhibits neurotransmitter release in the cortex but not in the stria- *Corresponding author. Tel: +81-44-244-7183; fax: +81-44-210-5893. E-mail address: kunio_torii@ajinomoto.com (K. Torii). Abbreviations: BOLD, blood oxygenation level-dependent; CBF, cere- bral blood flow; CC, cingulate cortex; DMH, dorsomedial hypothala- mus; fMRI, functional magnetic resonance imaging; Glu, L-glutamate; IC, insular cortex; LH, lateral hypothalamus; mPFC, medial prefrontal cortex; mPOA, medial preoptic area; MSG, monosodium L-glutamate; NTS, nucleus of the solitary tract; PVN, paraventricular nucleus; ROI, region of interests; SC, somatosensory cortex; TR, training period. Neuroscience 165 (2010) 244 –251 0306-4522/10 $ - see front matter © 2010 IBRO. Published by Elsevier Ltd. All rights reserved. doi:10.1016/j.neuroscience.2009.10.006 244