Hyperpolarization-activated cation channels are expressed in rat hypothalamic gonadotropin-releasing hormone (GnRH) neurons and immortalized GnRH neurons.
OBJECTIVE
The current research was conducted to determine whether hyperpolarization-activated cyclic nucleotide-gated (HCN1-4) channels are expressed in gonadotropin-releasing hormone (GnRH) neurons in the female rat hypothalamus and immortalized GnRH neurons (GT1-7 cells).


METHODS
Double-label fluorescence immunohistochemistry was used to colocalize HCN1-4 channels and GnRH in GnRH neurons in the female rat hypothalamus. Reverse transcriptase-polymerase chain reaction (RT-PCR), Western blotting, and immunocytochemistry were used to analyze HCN channel gene expression in GT1-7 cells.


RESULTS
Double-label fluorescence immunohistochemistry showed that 43% of hypothalamic GnRH neurons immunostained for HCN2 and 90% of GnRH neurons immunostained for HCN3. RT-PCR and Western blot showed expression of all four HCN channel subunits in GT1-7 cells. Double-label immunocytochemistry showed cytoplasmic immunostaining of HCN2 and HCN3 in GT1-7 cells.


CONCLUSIONS
This study demonstrates for the first time that HCN channels are expressed in GnRH neurons in the rat hypothalamus and GT1-7 cells. Our research supports the hypothesis that HCN channels may be involved in electrical bursting activity and pulsatile GnRH secretion in endogenous GnRH neurons and GT1-7 cells.
Arroyo
Armando
A
Department of Gynecology-Obstetrics, University at Buffalo, School of Medicine and Biomedical Sciences, Buffalo, New York 14222, USA. aarroyo2@buffalo.edu
Kim
Beomsu
B
Rasmusson
Randall L
RL
Bett
Glenna
G
Yeh
John
J
D000818:Animals; D015153:Blotting, Western; D002461:Cell Line, Transformed; D054815:Cyclic Nucleotide-Gated Cation Channels; D005260:Female; D005455:Fluorescent Antibody Technique; D015870:Gene Expression; D007987:Gonadotropin-Releasing Hormone / Q000378:metabolism*; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D007031:Hypothalamus / Q000166:cytology*; D007150:Immunohistochemistry; D007473:Ion Channels / Q000032:analysis / Q000235:genetics*; D009474:Neurons / Q000737:chemistry* / Q000502:physiology; D015221:Potassium Channels / Q000032:analysis / Q000235:genetics; D011301:Preoptic Area / Q000737:chemistry; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction
D054815:Cyclic Nucleotide-Gated Cation Channels; C513835:HCN3 protein, rat; C513836:HCN4 protein, rat; C578667:Hcn1 protein, rat; C578712:Hcn2 protein, rat; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D007473:Ion Channels; D015221:Potassium Channels; D012333:RNA, Messenger; D007987:Gonadotropin-Releasing Hormone