X-Message-Number: 24678
From: "Basie" <>
Subject: The effects of freezing on organ metabolism 
Date: Tue, 21 Sep 2004 00:54:53 -0400

ORGAN METABOLISM AND CRYOPROTECTANT SYNTHESIS IN SPRING PEEPERS Pseudacris
crucifer

Thomas A. Churchill and Kenneth B. Storey



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SUMMARY
The effects of freezing on organ metabolism were monitored over a 36 h time
course of freezing at -4 C in spring-collected specimens of a freeze
tolerant frog, the spring peeper Pseudacris crucifer. Within 2 min after
freezing began, glycogenolysis in liver had been activated as evidenced by
elevated levels of hexose phosphates and glucose in the organ. Freezing
stimulated a 3.3-fold increase in liver glycogen phosphorylase a activity
and a rapid phase of liver glucose production was maintained for at least
the first hour of freezing exposure (21  mol.g wet mass-1.h-1), followed by
a lower, steady rate of glucose increase of about 3  mol.gwm-1.h-1. Final
liver glucose concentration was 141  mol.gwm-1 after 36 h. Correlated with
this, changes in the concentrations of hexose phosphates and
fructose-1,6-bisphosphate in liver indicated that glucose production was
promoted by an inhibitory block on glycolysis at the phosphofructokinase
reaction during the rapid phase of glucose production. Other organs
accumulated glucose as a cryoprotectant during freezing exposure. Liver
energetics were not disrupted during the early phase of freezing but with
prolonged freezing (4-36 h) ATP content fell, ADP and AMP increased, and
energy charge ([ATP + ADP/2] / [ATP + ADP + AMP]) was reduced from 0.90 in
controls to 0.57 after 36 h freezing. Liver metabolism during freezing was
supported by fermentative reactions. Anaerobic glycolysis resulted in the
accumulation of both lactate and alanine and levels of fermentable free
amino acids (aspartate and glutamate) decreased significantly. Changes in
free amino acid patterns in skeletal muscle and heart indicated that these
also relied upon amino acid fermentation to help support energy metabolism
during freezing. In addition, both organs showed small increases in the
levels of numerous amino acids suggesting that freezing elevated
proteolysis. This study provides new evidence of the importance of amino
acids to anuran freezing survival and also demonstrates that the biochemical
mechanisms involved in regulating cryoprotectant output, previously
described only for the wood frog Rana sylvatica, may be general mechanisms
for regulating cryoprotectant pools in all freeze tolerant anurans

Churchill, T.A. and Storey, K.B. 1996. Organ metabolism and cryoprotectant
synthesis in spring peepers, Pseudacris crucifer. Copeia 196 (3) 517-525.

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