Brain Research 879 2000 29–41 www.elsevier.com locate bres
Research report
The role of intrinsic and agonist-activated conductances in determining the firing patterns of preoptic area neurons in the guinea pig
Edward J. Wagner , Cruz Reyes-Vazquez, Oline K. Rønnekleiv, Martin J. Kelly
Department of Physiology and Pharmacology , L334, Oregon Health Sciences University, 3181 S.W. Sam Jackson Park Road, Portland, OR 97201,
USA Accepted 11 July 2000
Abstract
Whole-cell and intracellular recordings were made in coronal hypothalamic slices prepared from ovariectomized female guinea pigs. 62 of preoptic area POA neurons fired action potentials in a bursting manner, and exhibited a significantly greater afterhyperpolariza-
tion AHP than did non-bursting POA neurons. The majority 70 of POA neurons n576 displayed a time-dependent inward rectification I that was blocked by CsCl 3 mM or by ZD 7288 30 mM. In addition, 51 of the cells expressed a low-threshold spike
h
LTS associated with a transient inward current I that was blocked by NiCl 200 mM. A smaller percentage of POA neurons 29
T 2
1
expressed a transient outward, A-type K current that was antagonized by a high concentration of 4-aminopyridine 3 mM. Moreover,
POA neurons responded to bath application of the m-opioid receptor agonist DAMGO 93 or the GABA receptor agonist baclofen
B
83 with a membrane hyperpolarization or an outward current. These responses were accompanied by a decrease in input resistance or an increase in conductance, respectively, and were attenuated by BaCl 100 mM. In addition, the reversal potential for these responses
2 1
closely approximated the Nernst equilibrium potential for K . These results suggest that POA neurons endogenously express to varying
1
degrees an AHP, an I , an I and an A-type K current. The vast majority of these neurons also are inhibited upon m-opioid or GABA
h T
B 1
receptor stimulation via the activation of an inwardly-rectifying K conductance. Such intrinsic and transmitter-activated conductances
likely serve as important determinants of the firing patterns of POA neurons.
2000 Elsevier Science B.V. All rights reserved.
Theme : Excitable membranes and synaptic transmission
Topic : Postsynaptic mechanisms
Keywords : Preoptic area; Firing; Afterhyperpolarization; Inward rectification; Calcium current
1. Introduction thermoregulation, stress, reproduction and maternal be-
havior [3,20,34,36]. The preoptic area POA of the hypothalamus is an
Thermoregulatory neurons are found in the POA that integrative center for many important physiological func-
respond to changes in body temperature [2–4,50]. Warm- tions. It receives input from many different brain regions,
sensitive neurons increase their firing rate in response to including hypothalamic and other limbic areas, cortex,
increases in body temperature, thereby promoting heat midbrain and brain stem [34,35,43]. Likewise, the POA
dissipation from the body. In contrast, cold-sensitive sends neuronal projections to many of these same regions
neurons become activated upon a decrease in body tem- [34,44,49]. Some of the inputs to and neuronal projections
perature, which facilitates heat production and retention. from the POA appear to be reciprocal [49]. It is therefore
While intrinsic conductances are undoubtedly important not surprising that the POA plays an important role in
determinants of the firing patterns of thermosensitive many physiological processes including, but not limited to,
neurons, these cells are also modulated by pyrogens i.e., interleukin-6, opioids and g-aminobutyric acid GABA
B
receptor agonists,
all of
which elicit
hyperthermia [8,50,51]. The increase in body temperature caused by
Corresponding author. Tel.: 11-503-494-5830; fax: 11-503-494-
interleukin-6 arises from an inhibition of warm-sensitive
4352. E-mail address
: wagnereohsu.edu E.J. Wagner.
neurons and an activation of cold-sensitive neurons, both
0006-8993 00 – see front matter
2000 Elsevier Science B.V. All rights reserved. P I I : S 0 0 0 6 - 8 9 9 3 0 0 0 2 6 9 8 - 6
30 E
of which are blocked by the opioid receptor antagonist 2.2. Drugs
naloxone [50]. Thus, the hyperthermia evoked by these factors is the result of a decreased capacity to dissipate
All drugs were initially dissolved in Milli-Q H 0 unless
2
heat, and an enhanced ability to produce and retain heat. otherwise stated. TTX Sigma Chemical Co., St. Louis,
The POA also contains gonadotropin hormone-releasing MO, USA was dissolved in Milli-Q H O and diluted to
2
hormone GnRH neurons that project to the median the appropriate volume with 0.1 acetic acid final
eminence, and the neuropeptide released from these neu- concentration 1 mM; pH 4–5. Cesium chloride CsCl;
rons acts as the primary secretagogue in the release of J.T. Baker Inc., Phillipsburg, NJ, USA was prepared as a
gonadotropins i.e., follicle stimulating hormone, luteiniz- stock solution of 0.2 M. N-ethyl-N-phenylamino-1,2-di-
ing hormone LH from the anterior pituitary [15,42]. methyl-6-methylamino pyrimidinium chloride ZD 7288;
POA neurons receive inhibitory GABAergic input [28], as Tocris Cookson Inc., Ballwin, MO, USA was prepared as
well as synaptic input from b-endorphin neurons emanat- a stock solution of 10 mM. Nickel chloride hexahydrate
ing from the arcuate nucleus [9,18]. Activation of GABA NiCl ; Sigma was prepared as a stock solution of 0.1 M.
B 2
receptors inhibits naloxone-stimulated LH release in a 4-Aminopyridine 4-AP; Aldrich Chemical Co., Mil-
variety of experimental models [7,11,30], thereby indicat- waukee, WI, USA was prepared as a stock solution of 0.1
ing an indirect opioid modulation of LH secretion through M. Tetraethylammonium TEA; Sigma was prepared as a
an interaction with GABAergic neurons. There is, how- stock solution of 10 mM. DAMGO Peninsula Laborator-
ever, ample evidence to suggest that opioid peptides and ies Inc., Belmont, CA, USA and met-enkephalin Penin-
GABAergic ligands affect changes in reproductive status sula were prepared as stock solutions of 1 and 5 mM,
by acting at the level of the POA to directly inhibit GnRH respectively. 6-Baclofen 6-b-aminomethyl-4-chlo-
and thereby LH secretion [25,28,45]. robenzenepropanoic acid; Sigma was dissolved in 0.1 N
The purpose of the present study was to characterize the HCl to a stock concentration of 40 mM. Barium chloride
prominent intrinsic and inhibitory, agonist-activated con- dihydrate BaCl ; Sigma was prepared as a stock solution
2
ductances in POA neurons of the ovariectomized female of 10 mM. Aliquots of the various stock solutions were
guinea pig. To this end, whole-cell and intracellular stored at 2808C TTX, DAMGO, met-enkephalin, ba-
recordings were made from coronal hypothalamic slices. clofen, 2208C ZD 7288or 48C CsCl , NiCl , 4-AP,
2 2
2 4
Methionine met-enkephalin and [
D
-Ala , N-Me-Phe , TEA, BaCl until used for experimentation.
2 5
Gly-ol ]-enkephalin DAMGO, opioid peptides selective for the m-receptor [17], were used to assess the effects of
2.3. Hypothalamic slice preparation m-opioid receptor activation. Likewise, the selective
GABA receptor agonist baclofen [6] was used to evaluate On the day of experimentation, the animal was decapi-
B
the effects of GABA receptor activation. The results are
tated, and its brain rapidly removed from the skull. The
B
discussed in a context pertinent to the important role of the brain was rinsed with ice-cold artificial cerebrospinal fluid
POA in physiological processes such as thermoregulation aCSF; NaCl, 124; KCl, 5; NaH PO , 2.6; dextrose, 10;
2 4
and reproduction. HEPES, 10; MgSO , 2; CaCl , 2; in mM and the
4 2
hypothalamus immediately dissected. Four coronal slices 450 mM through the rostro-caudal extent of the POA
were cut with the aid of a vibratome. The slices ranged
2. Materials and methods from the organum vasculosum laminae terminalis at the
