Attenuation of the Counter-Regulatory Glucose Response in CVLM C1 Neurons: A Possible Explanation for Anorexia of Aging

Abstract

This study aimed to determine the effect of age on CVLM C1 neuron glucoregulatory proteins in the feeding pathway. Male Sprague Dawley rats aged 3 months and 24 months old were divided into two subgroups: the treatment group with 2-deoxy-d-glucose (2DG) and the control group. Rat brains were dissected to obtain the CVLM region of the brainstem. Western blot was used to determine protein expression of tyrosine hydroxylase (TH), phosphorylated TH at Serine40 (pSer40TH), AMP-activated protein kinase (AMPK), phosphorylated AMPK (phospho AMPK), and neuropeptide Y Y5 receptors (NPY5R) in CVLM samples. Immunofluorescence was used to determine TH-, AMPK-, and NPY5R-like immunoreactivities among other brain coronal sections. Results obtained denote a decrease in basal TH phosphorylation levels and AMPK proteins and an increase in TH proteins among aged CVLM neurons. Increases in the basal immunoreactivity of TH+, AMPK+, NPY5R+, TH+/AMPK+, and TH+/NPY5R+ were also observed among old rats. Young treatment-group rats saw a decrease in TH phosphorylation and AMPK proteins following 2DG administration, while an increase in AMPK phosphorylation and a decrease in TH proteins were found among the old-treatment-group rats. These findings suggest the participation of CVLM C1 neurons in counter-regulatory responses among young and old rats. Altering protein changes in aged CVLM C1 neurons may attenuate responses to glucoprivation, thus explaining the decline in food intake among the elderly.

Keywords: anorexia of aging; catecholaminergic neurons; caudal ventrolateral medulla; feeding response; glucoprivation.

Figure 1

Distribution of TH-containing neurons and colocalization with AMPK or NPY5R in CVLM in different age groups. The highlighted area in the box indicates the CVLM region (A). TH co-localized with AMPK (B) and NPY5R (C) in CVLM neurons. The scale bars are 200 and 100 μm.

Figure 2

Effects of age on the distribution of TH, AMPK, and NPY5R in CVLM neurons. The numbers of neurons containing TH, AMPK, and NPY5R in CVLM were compared between age groups (A). Further analysis was conducted to show the colocalization of TH+/AMPK+ and TH+/NPY5R+ in the same brain region (B). Each bar and associated error bar represent the mean value of neuronal number ±SEM from one experiment (n = 3 per group). *** (p < 0.001) represents a significant difference between control groups of young rats and old rats, respectively.

Figure 3

Effects of age and glucoprivation condition on the level of phosphorylated AMPK (A), total AMPK (tAMPK) (B), pSer40 TH (C), TH (D), and NPY5R (E). Each bar and associated error bar represents mean value of the intensity band ± SEM from one experiment (n = 6 per group). Representative Western blots (A–D) depicting effects of age and 2DG or saline; each lane represents a single animal. ++ (p < 0.01) and + (p < 0.05) represent significant differences between control groups of young and old rats, respectively, and ** (p < 0.01) and * (p < 0.05) represent significant differences between 2DG and control groups of the same-aged rats.

Figure 4

Summary of the glucose counterregulatory responses in CVLM neurons following glucoprivation in young and aged rats [56].