 |
Awarded by ATS at
|
Be sure to check out the cover article contributed by Dr. Pang
in
our Targeting Trends newsletter (Second Quarter,
2009).
865.23/MM24 Stimulus-, circuit- and intracellular-level determinants
of MAP kinase and CREB activation in parvicellular hypothalamic paraventricular
neurons
A M Khan, K L Rapp, T A Ponzio, G Sanchez-watts, A G Watts
featuring IT-03 Anti-DBH-SAP (Poster;
Wednesday, Nov 19, 2008, 3:00 PM - 4:00 PM)
Systemic insulin or 2-deoxyglucose (2-DG) rapidly elevate phosphorylated MAP
kinases (phospho-ERK1/2) and/or CRH hnRNA in PVHp neurons, and increase circulating
ACTH and corticosterone. These neuroendocrine responses are likely driven by
hindbrain-originating catecholaminergic (CA) neuron subpopulations, which richly
innervate the PVHp and are activated by glycemic challenges. Supporting this,
acute in vivo or in vitro PVH delivery of the prototypical catecholamine, norepinephrine
(NE), recapitulates these responses (J Neurosci, 2007, 27:7344-7360). Here,
we determined whether PVHp ERK/CREB phosphorylation responses require: 1) intact
CA afferents, when triggered by three distinct in vivo challenges; and 2) upstream
MEK kinase activity, when triggered by NE application in acute hypothalamic
slices maintained in vitro.
Methods. Rats given PVH microinjections of anti-dopamine-b-hydroxylase (DBH)-saporin
antibody-toxin conjugate (DSAP) or mIgG-saporin control conjugate received
either normal 0.9% saline vehicle or one of three systemic challenges: insulin
(2 U/kg, i.v.); 2-DG (250 mg/kg, i.v.); or hypertonic saline (1.5 M, i.p.)
and sacrificed 30 min later. Brains were processed for CRH mRNA/hnRNA hybridization,
or DBH, phospho-ERK or phospho-CREB immunocytochemistry. Plasma was collected
for hormone determinations at 0 and 30 min. In separate in vitro studies, acute
hypothalamic slices received either no treatment (controls), or received bath-applied
NE (100 mM) in the presence or absence of the MEK inhibitor, U0126 (10 mM),
or the inactive MEK inhibitor analogue, U0124 (10 mM). Ten min later, slices
were placed in fixative.
Results. 1. Sham-lesioned animals: Relative to vehicle, all challenges elevated
phospho-ERK1/2, phospho-CREB, and ACTH/corticosterone levels; and, except for
insulin, also increased CRH hnRNA. 2. Lesioned animals: DSAP treatment selectively
destroyed hindbrain-originating CA afferents. In insulin- and, to a lesser
extent, 2-DG-treated animals, this loss was accompanied by markedly reduced
PVH phospho-ERK1/2 and circulating ACTH/corticosterone. In contrast, these
responses remained robust in CA-deafferented hypertonic saline-treated rats.
Phospho-CREB levels were differentially reduced relative to phospho-ERK in
lesioned rats. 3. Slices: NE-induced PVH elevations of phosphorylated ERK1/2
and CREB were reduced markedly by U0126, but not U0124, pre-treatment.
Conclusions. PVHp phospho-ERK selectively couples to CA afferents during glycemic
challenges and ERK/CREB recruitment appears to require MEK activity.
