Molecular analysis of vasopressin receptors in the rat nephron. Evidence for alternative splicing of the V2 receptor

Abstract

Expression and regulation of vasopressin V2 and V1a receptors were studied at the mRNA level in the rat kidney. Two V2 mRNA variants were identified and shown to arise from a single gene by alternative splicing using one donor and two different acceptor sites. The long (V2L) form encodes the adenylyl cyclase-coupled receptor. The short (V2S) form lacks the nucleotide sequence encoding the putative seventh transmembrane domain and undergoes a frame shift in its 3'end coding region; it is inactive on the cyclase pathway in transfected cells. Measurement of mRNAs, carried out by quantitative reverse transcription-polymerase chain reaction (RT-PCR) on microdissected nephrons, demonstrated that neither V2L, V2S nor V1A mRNAs are expressed in glomeruli and proximal tubules (< 100 mRNA copies/glomerulus or mm of tubular length), whereas they are present in the ascending limb of Henle's loop and in the collecting tubule. The V2L mRNA, which is always predominant in these structures, is expressed throughout the collecting tubule at 10 times higher levels (30,000 copies/mm) than in the thin and thick ascending limbs. The ratio of the V2S over V2L mRNA is constant (15%) in all nephron segments; hence high V2S levels are only observed in the collecting tubule. The V1A mRNA is slightly expressed in the thin ascending limb, absent in the thick ascending limb and reaches its maximum in the cortical collecting duct (4,000 copies/mm), before gradually decreasing to undetectable levels in the terminal collecting duct. Finally, in vivo administration of a vasopressin V2 agonist decreased by 50% V2L and V2S mRNAs, but did not alter the V1A mRNA level. We conclude that this study provides the quantitation, on a molar basis, of vasopressin receptor mRNAs in kidney tubules and demonstrates the occurrence of two V2 mRNA spliced variants which are similarly down-regulated.

Molecular clock is involved in predictive circadian adjustment of renal function

“…Microdissection of DCT/CNT or CCD was performed from collagenase-treated kidneys as described previously (32).…”

Downregulation of the V2 vasopressin receptor in dehydration: mechanisms and role of renal prostaglandin synthesis

“…Specific primers for the V2 receptor and AQP2 were designed. The sequence of sense and antisense V2 receptor primers were defined by bases 349 -368 (5Ј-TACCTGCAGATGGTGGGCAT-3Ј) and bases 910 -929 (5Ј-AGCAACACAAAGGGGGGTCT-3Ј), respectively (9,20). This corresponds the long form of V2 receptor mRNA in Firsov's classification (9).…”

“…V1a receptors mediate the vasopressor effects of AVP and are localized in renal vessels and in the thick ascending limbs and the collecting ducts (26,28,34). In contrast, V2 receptors mediate the antidiuretic action of AVP and are localized in the thick ascending limbs and the collecting ducts (9,16,25,34). In the collecting ducts, the V2 receptor is located in the basolateral membrane of principal cells, while V1a receptor is mainly in the luminal membrane of both principal and intercalated cells (9,25,33).…”

“…In contrast, V2 receptors mediate the antidiuretic action of AVP and are localized in the thick ascending limbs and the collecting ducts (9,16,25,34). In the collecting ducts, the V2 receptor is located in the basolateral membrane of principal cells, while V1a receptor is mainly in the luminal membrane of both principal and intercalated cells (9,25,33). Urine concentration is caused by the binding of AVP to the V2 receptor and by the subsequent activation of aquaporin-2 (AQP2) (13,14,19,23).…”

Increased Renal Responsiveness to Vasopressin and Enhanced V2 Receptor Signaling in RGS2 −/− Mice

“…Microdissection of different parts of the nephron was performed from collagenase-treated kidneys as described previously (11).…”