Eating in mice with gastric bypass surgery causes exaggerated activation of brainstem anorexia circuit

Abstract

Background/objective: Obesity and metabolic diseases are at an alarming level globally and increasingly affect children and adolescents. Gastric bypass and other bariatric surgeries have proven remarkably successful and are increasingly performed worldwide. Reduced desire to eat and changes in eating behavior and food choice account for most of the initial weight loss and diabetes remission after surgery, but the underlying mechanisms of altered gut-brain communication are unknown.

Subjects/methods: To explore the potential involvement of a powerful brainstem anorexia pathway centered around the lateral parabrachial nucleus (lPBN), we measured meal-induced neuronal activation by means of c-Fos immunohistochemistry in a new high-fat diet-induced obese mouse model of Roux-en-Y gastric bypass (RYGB) at 10 and 40 days after RYGB or sham surgery.

Results: Voluntary ingestion of a meal 10 days after RYGB, but not after sham surgery, strongly and selectively activates calcitonin gene-related peptide neurons in the external lPBN as well as neurons in the nucleus tractus solitarius, area postrema and medial amygdala. At 40 days after surgery, meal-induced activation in all these areas was greatly diminished and did not reach statistical significance.

Conclusions: The neural activation pattern and dynamics suggest a role of the brainstem anorexia pathway in the early effects of RYGB on meal size and food intake that may lead to adaptive neural and behavioral changes involved in the control of food intake and body weight at a lower level. However, selective inhibition of this pathway will be required for a more causal implication.

Fig. 1

Effects of Roux-en-Y gastric bypass surgery on body weight (a), body composition (b), and food intake (c) in mice made obese by exposure for 14 weeks to a two-choice diet consisting of high-fat and regular (low-fat) chow. a: Change in body weight after RYGB (filled circles; n = 39 for days 1–8, n = 15 for days 10–40) or sham surgery (open circles; n = 34, 16) expressed in percent of pre-surgical body weight. Double arrows indicate the time of meal tests at 10 and 40 days after surgery. b: Body weight, absolute fat mass, relative fat mass (adiposity), and lean mass before and 40 days after RYGB (n = 15) or sham surgery (n = 16). Bars that do not share the same letters are significantly different from each other (based on ANOVA followed by Bonferroni-corrected multiple comparisons). c: Total food intake before and after RYGB (filled circles; n = 28 for days 1–8, n = 4 for days 10–36) or sham-surgery (open circles; n = 24, 7). The two horizontal bars mark periods during which chow preference (based on calories) was measured in both groups (data shown in inset). * p<0.05, RYGB vs. Sham.

Fig. 2

Meal-induced c-Fos expression in the lateral parabrachial nucleus of mice with RYGB or sham surgery. a, b: Representative micrographs showing c-Fos expression in the external lateral parabrachial nucleus in a mouse with sham-surgery (a) or RYGB (b) after consumption of a meal at 10 days after surgery. CGRP expression (b′), merged c-Fos (black) and CGRP (pink) expression (b″), as well as an enlarged detail of double-labeling (b‴) are also shown for the RYGB mouse. c: High fat test meal size was not significantly different between RYGB and sham groups at either 10 or 40 days. d,e: Quantitative analysis of the number of c-Fos positive cells (d) and percent of CGRP positive cells expressing c-Fos (e) for meal consumption tests 10 days after surgery. f: Quantitative analysis of the number of c-Fos positive cells for meal consumption tests 40 days after surgery. Bars that do not share the same letters are significantly different from each other (based on ANOVA followed by Bonferroni-corrected multiple comparisons). Means ± SEM of 5–10 mice. Scale bar in a: 100 μm for a–b″ and 30 μm for b‴. Abbreviations: el, external lateral subnucleus; KF, Kölliker-Fuse nucleus; scp, superior cerebellar peduncle; vl, ventrolateral subnucleus.

Fig. 3

Meal-induced c-Fos expression in the dorsal vagal complex of mice with RYGB or sham surgery. a, b: Representative micrographs showing c-Fos expression in the NTS and AP in a mouse with sham (a) or RYGB (b) surgery 10 days after surgery. TH expression (b′) and merged c-Fos (black) and TH (pink) expression (b″), as well as an enlarged detail of double-labeling (b‴) are also shown for the RYGB mouse (white arrow indicates lonely double-labelled neuron). c–f: Quantitative analysis of the number of c-Fos positive cells in the NTS 10 days (c) and 40 days (f) after surgery, as well as number (d) and percent (e) of TH neurons also expressing c-Fos for 10 day meal consumption tests. g–j: Quantitative analysis of the number of c-Fos positive cells in the area postrema 10 days (g) and 40 days (j) after surgery, as well as number (h) and percent (i) of TH neurons also expressing c-Fos for 10 day meal consumption tests. Means + SEM of 4–10 mice. Bars that do not share the same letters are significantly different from each other (based on ANOVA followed by Bonferroni-corrected multiple comparisons). Scale bar in a: 100 μm for a–b″ and 25 μm for b‴.

Fig. 4

Meal-induced c-Fos expression in the central nucleus of the amygdala (CeA) of mice with RYGB or sham surgery. a, b: Representative micrographs showing meal-induced c-Fos expression in the lateral part of CeA, 10 days after sham (a) or RYGB surgery (b). c, c′: Low (c) and higher power (c′) images of double-labeled section showing proximity of meal-induced c-fos (black) and CGRP-immunoreactive projections (pink) in CeA of mouse 10 days after RYGB. d, e: Quantitative analysis of meal-induced c-Fos expression in the CeA at 10 (c) and 40 (d) days after RYGB or sham surgery. Means + SEM of 4–8 mice. Bars that do not share the same letters are significantly different from each other (based on ANOVA followed by Bonferroni-corrected multiple comparisons). Scale bar in a: 100 μm for a–c; 25 μm for c′. Abbreviations: BLA, anterior basolateral amygdaloid nucleus; CeA, central amygdaloid nucleus; I, intercalated nuclei amygdala; ic, internal capsule; LGP, lateral globus pallidus.

Fig. 5

Correlations between meal size and c-Fos expression in the lPBN (a,b), NTS (c), and CeA (d) of mice with RYGB (n = 6–8; upper panels) or sham surgery (n = 3–9; lower panels) 10 days after surgery. For the lPBN, the correlation between meal size and percent of activated CGRP⁺ neurons is also shown (b). All correlation coefficients and p values refer to meal tests 10 days after surgery. Correlations 40 days after surgery and in sham animals at both time points were weak and non-significant.