Targeted disruption of the gene for pituitary adenylate cyclase-activating polypeptide (PACAP) in mouse results in metabolic dysfunction.
Date
2018-11-07
Authors
Gray, Sarah Louise
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Abstract
A recently discovered peptide hormone, pituitary adenylate cyclase-activating
polypeptide (PACAP) regulates several endocrine systems affecting essential
physiological processes such as metabolism, growth, reproduction, and the stress
response. PACAP acts as a hypophysiotropic factor, is a potent secretogogue of insulin,
regulates production and release of catecholamines from the adrenal medulla and acts as
a neuromodulator in the sympathetic and parasympathetic nervous system. The primary
structure of PACAP has been highly conserved during the evolution of chordates
suggesting it plays an important physiological role. The objective of my thesis was to
identify PACAP’s primary physiological function and to determine if it is essential for
survival by generating a mouse line deficient in PACAP through targeted disruption of
the PACAP gene locus.
Postnatal PACAP expression was examined to determine sites of peripheral
PACAP production. In addition, several splice variants of the PACAP gene with
alternate 5’untranslated regions were identified suggesting a complex system for
regulating expression of the mouse PACAP gene.
A targeting vector that allows tissue specific or developmental stage specific
knockout of the PACAP gene was constructed in the event that PACAP gene deletion
resulted in embryonic lethality. PACAP null mice were generated from homologously
recombined embryonic stem cells. Initial characterization of the PACAP null mice
determined that in the absence of PACAP, mice died within the first two postnatal weeks
with abnormal lipid metabolism. Lipid accumulation was present in liver, heart and
skeletal muscle and serum lipids were high. Mitochondrial dysfunction in the liver was
not the cause of the lipid accumulation, as P-oxidative function was normal. I conclude
that PACAP null mice are unable to regulate lipid release from white adipose tissue
stores, resulting in a flood of lipids to non-adipose tissues.
The abnormal distribution of lipids observed in the PACAP null mice is
characteristic of diabetes type 2, yet classical insulin resistance is not observed. Thus,
elevated insulin levels were accompanied by low blood glucose levels and the response to
a glucose challenge was normal. The uncontrolled release of free fatty acids may result if
glucose that is taken up by cells can not be utilized and an alternate energy source is
required or if white adipocytes only are insulin resistant.
The PACAP null mice were temperature sensitive, in that when raised at 21“C
they exhibited metabolic dysfunction and died by two weeks of age. At 24°C most (85%)
of the mice survived to adulthood with no obvious signs of metabolic dysfunction. We
have determined that the inability of the PACAP null pups to thermoregulate normally
when exposed to a lower environmental temperature may be associated with decreased
norepinephrine levels to the brown adipose tissue. PACAP may be important for the
production and release of catecholamines in the adrenal gland or within the sympathetic
nervous system in times of prolonged stress.
A mechanistic connection between the lipid abnormalities and the temperature
sensitivity in the PACAP null pups has yet to be made. Catecholamines affect a wide
range of tissues and the problems associated with insulin regulation within the PACAP
null mice may be due to the imbalance in catecholamine production. As one of two main
stress response systems, the sympathetic nervous system elicits a vital coping mechanism
in times of stress and PACAP’s ability to regulate this system may explain why the
primary structure of PACAP has remained so highly conserved. PACAP is a wide acting
hormone and therefore the metabolic problems seen in the PACAP null mice may result
from altered regulation of several endocrine systems at once. Targeted disruption of the
PACAP gene in mouse has revealed a role for PACAP in the regulation of lipid
metabolism and in the sympathetic control of thermoregulation.
Description
Keywords
Hormones, Physiological effect, Endocrine genetics