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. 2000 Nov;47(5):653–660. doi: 10.1136/gut.47.5.653

Restricted diet rescues rat enteric motor neurones from age related cell death

T Cowen 1, R Johnson 1, V Soubeyre 1, R Santer 1
PMCID: PMC1728112  PMID: 11034581

Abstract

BACKGROUND—Alone among autonomic neurones, enteric neurones are known to be vulnerable to age related cell death; over 50% may be lost in aging rodents. A previous study demonstrated unexpectedly that neurones of the myenteric plexus from rats fed a restricted diet appeared not to suffer from extensive cell death in contrast with previous studies of ad libitum fed animals.
AIMS—To compare myenteric neurone numbers in the ileum of young and aging male Sprague-Dawley rats fed either ad libitum or a restricted diet.
METHODS—Neurones were counted in whole mount preparations of rat ileum stained immunohistochemically for the pan-neuronal marker PGP9.5, for choline acetyltransferase, or for nitric oxide synthase, or with NADH or NADPH histochemistry.
RESULTS—Neurone numbers in the rat myenteric plexus were substantially affected by the dietary regimen: ad libitum feeding (50-60 g per day of standard rat chow) resulted in the death of about 50% of myenteric neurones in 24 month Sprague-Dawley rats, while numbers were unchanged when the daily dietary intake was halved between the ages of six and 24 months. Animals fed a double restricted diet (15 g per day) showed no cell loss at 30 months, as well as the predicted increase in longevity. Neurone loss was largely complete by 16 months in ad libitum fed animals. Numbers of cholinergic (possibly motor) neurones, as demonstrated by choline acetyltransferase immunohistochemistry, were substantially reduced in ad libitum fed aging rats but not in animals fed a restricted diet. Loss of cholinergic neurones after ad libitum feeding was confirmed by reduced numbers of neurones of a size range matching that of cholinergic neurones.
CONCLUSIONS—Ad libitum feeding of adult rats has adverse effects on the survival of myenteric neurones, neurone loss commencing before 16 months of age. Cholinergic neurones appear to be particularly vulnerable to the effects of diet. Restricting dietary intake from six months of age prevents neurone loss almost entirely up to 30 months of age in these rats.


Keywords: restricted diet; enteric neurones; aging; neuronal cell death

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Figure 1  .

Figure 1  

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Photomicrographs showing PGP immunoreactive neurones in the myenteric plexus of young (4-6 month (A)), ad libitum (AL) fed (AL 16 month (B, C), AL 24 month (D)), and restricted diet (RD 24 month (E) and double RD 30 month (F)) rats. Note the ganglion full of brightly stained neurones in (A) (large arrows), and also in (B), (E), and (F). The secondary (medium arrows) and tertiary (small arrows) plexuses are also seen in all preparations. Nerve loss is clearly seen as spaces within the majority of ganglia in AL 16 month and AL 24 month animals (C, D) (arrows) but not in all AL animals (B). Scale bar 50 µm.

Figure 2  .

Figure 2  

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Histogram showing the number of PGP immunoreactive neurones in the myenteric plexus of young (4-6 month) rats, ad libitum fed (AL) rats at 16, 20, and 24 months, rats fed a restricted diet (RD) at 24 months, and rats fed a double restricted diet (DRD) at 30 months. Note the comparable loss of neurones in AL 16, AL 20, and AL 24 month groups compared with young, RD 24 month and DRD 30 month groups. These losses were highly significant (see table 2), reaching >50% in the AL groups. Also, note the lack of cell loss in senile (DRD 30 months) animals fed a double restricted diet.

Figure 3  .

Figure 3  

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Photomicrographs showing NADH histochemical staining in the myenteric plexus of young 4-6 month (A), old ad libitum fed (AL 24 month) (B), and restricted diet (RD 24 month) (C) rats. Note the reduced density of neurones in ganglia from AL (B) as well as RD (C) rats, indicating that NADH is not suitable as a pan-neuronal marker (see text and table 1). Also, note the increased size of neurones in (B) and (C). Scale bar 50 µm.

Figure 4  .

Figure 4  

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Photomicrographs showing NADPH histochemical staining for nitrergic neurones in ganglia from the myenteric plexus of young (A) and ad libitum fed 24 month (B) rats. Note the comparable density of stained neurones. Scale bar 50 µm.

Figure 5  .

Figure 5  

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Histogram showing size (average diameter) distributions of neurones in different age groups following ad libitum (AL) and restricted diet (RD) feeding in 24 month old rats. Note the rightward shift of neurone sizes at all ages after six months, indicating continued neurone growth in adult life. The frequency of neurones of 15-17.5 µm diameter is reduced in the 24 month AL group compared with the 24 month RD group, matching the average size of cholinergic neurones measured separately in neurones from 24 month RD rats (data not shown).

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