Detection of thromboxane A(2) receptor mRNA in rabbit nodose ganglion neurons.
Thromboxane A(2) (TXA(2)) is an arachidonic acid metabolite that is released during tissue trauma and elicits platelet aggregation and vascular smooth muscle contraction. Previous research has shown that TXA(2) stimulates pulmonary and cardiac vagal afferent neurons. Therefore, we hypothesized that the presence of the TXA(2) receptor (TP) in vagal neurons would allow for stimulation or modulation of these neurons by TXA(2). To test this hypothesis, single cell RT-PCR was employed using neurons obtained from primary cell cultures of nodose ganglia excised from adult rabbits. Since the sequence for the rabbit TP gene was unknown, a portion of the rabbit TP cDNA was first amplified, cloned, and sequenced. Primer sets for TP were then designed based on this sequence and used in conjunction with a neuronal marker, medium weight neurofilament (NFM), in multiplex RT-PCR reactions. Ninety-three cells were isolated from culture and RT-PCR was carried out on individual cells. Using an aliquot from the initial RT-PCR reaction, a second round of PCR was then employed in which the NFM and TP primer sets were split up into separate reactions. Twenty-three of the 82 cells that were positive for NFM were also positive for TP. Therefore, we conclude that the presence of TP mRNA in a subset of cultured nodose ganglion neurons allows for the possibility that TXA(2) may directly stimulate or modulate vagal afferent neurons.
Wacker
Michael J
MJ
Department of Molecular Biosciences, University of Kansas, 1200 Sunnyside Avenue, Lawrence, KS 66045, USA. mjwacker@ku.edu
Tyburski
John B
JB
Ammar
Chad P
CP
Adams
M Christopher
MC
Orr
James A
JA
D000818:Animals; D001483:Base Sequence; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D009474:Neurons / Q000378:metabolism*; D009620:Nodose Ganglion / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D011817:Rabbits; D044244:Receptors, Thromboxane A2, Prostaglandin H2 / Q000235:genetics* / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012689:Sequence Homology, Nucleic Acid
D012333:RNA, Messenger; D044244:Receptors, Thromboxane A2, Prostaglandin H2