Co‐ordinated expression of 5‐HT_2C receptors with the NCX1 Na⁺/Ca²⁺ exchanger in histaminergic neurones

Abstract

The different roles of Na + /Ca 2+ (NCX) exchangers and Na + / Ca 2+ /K + (NCKX) exchangers in regulation of the ionic homeostasis in neurones are poorly understood. We have previously shown that serotonin excites histaminergic tuberomamillary (TM) neurones by activation of 5-HT 2C -receptors and Na + /Ca 2+ exchange. With the help of single-cell RT-PCR (sc-RT-PCR) we have now determined the coexpression pattern of different subtypes of NCX and NCKX with serotonin receptors. The majority of TM neurones express NCX1, NCX2 and NCKX3. Serotonin 2C receptor-mRNA was detected in 70% while 5-HT 2A mRNA was found in only 10% of TM neurones. In all neurones expressing the 5-HT 2C receptor NCX1-mRNA was present. Double immunostaining revealed the presence of the NCX1 protein in histidine decarboxylasepositive neurones. In the majority of TM neurones one or two out of five isoforms, NCX1.4, NCX1.5, NCX1.7, NCX1.14, NCX1.15, were detected by cDNA sequencing and/or by restriction analysis. The alternative splicing region is important for the Ca 2+ sensitivity and presumably for the modulation of NCX1 function by second messengers. We conclude that several exchanger-subtypes can be coexpressed in single neurones and that TM cells are heterogeneous with respect to their calcium homeostasis regulation. Calcium plays a central role in neuronal signalling and after a quick rise in response to physiological stimuli free calcium must be immediately removed and finally extruded from the cell in order to maintain its low resting level. Two systems, the plasma membrane Ca 2+ ATPase (Carafoli 1994) and the sodium-calcium exchangers (Philipson and Nicoll 1992) are responsible for the efflux of intracellular calcium. These exchangers come from two gene families: Na + /Ca 2+ (NCX) and Na + /Ca 2+ /K + (NCKX) exchangers (Schulze et al. 2002b). In vitro expression of each known gene results in the formation of a functional protein which imports across the cellular membrane 3Na + for every Ca 2+ that is pumped out (NCX; Kimura et al. 1987) or 4Na + for 1K + plus 1Ca 2+ (NCKX; Blaustein and Lederer 1999). Function of these exchangers in native neurones remains elusive mainly because of the lack of selective antagonists for NCX versus NCKX. Nevertheless, recent studies attribute an important role to NCKX and NCX for Ca 2+ extrusion from living neurones (Ranciat-McComb et al. 2000;Lee et al. 2002).We have recently shown that serotonin depolarizes histaminergic neurones and increases their firing rate (Eriksson et al. 2001b. This action was suppressed by the two Na + /Ca 2+ exchange inhibitors KB R7943 (80 lM, 2-[2-[4-(4-nitrobenzyloxy)phenyl]ethyl]isothiourea methanesulphonate) and DCB (100 lM, 3¢,4¢-dichlorobenzamil). Moreover, other characteristic features of serotonin-induced depolarization, such as dependence on external Na + and temperature suggested involvement of an exchanger. However, the available pharmacological tools were inadequate for an unambiguous conclusion on the type of Na + /Ca 2+ exchanger involved in the observed p...

International Union of Basic and Clinical Pharmacology. XCVIII. Histamine Receptors

“…This leads to calcium ion entry through calcium channels, cation channels of the transient receptor potential channel type Doreulee et al, 2003), and stimulation of a Na + /Ca 2+ exchanger Sergeeva et al, 2003). Nicotinic acid adenine dinucleotide phosphate (NAADP) has also been suggested as a potential second messenger in histamine-induced Ca 2+ release from lysosome-like acidic compartments, functionally coupled to the endoplasmic reticulum via H 1 receptor in endothelial cells.…”

Pharmacology of Brain Na+/Ca2+Exchanger: From Molecular Biology to Therapeutic Perspectives

“…Thus, it has been demonstrated that the serotonin-induced increase of the firing rate of histaminergic tuberomammilar neurons is weakened by NCX inhibitors (Eriksson et al, 2001a,b). Interestingly, the serotonin receptor 5-HT 2C , which mediates this effect on histaminergic tuberomammillar neurons, is also coexpressed with NCX1 (Sergeeva et al, 2003). Moreover, in the same histaminergic neurons, NCX activity plays a relevant role in orexin-induced depolarization (Eriksson et al, 2001b).…”

Editing of AMPA and Serotonin 2C Receptors in Individual Central Neurons, Controlling Wakefulness

“…We have correlated ADAR-, 5-HT 2C R -and GluR-expression using the single-cell RT-PCR technique in individual neurons. We selected for this purpose hypothalamic histaminergic neurons acutely isolated from the tuberomamillary nucleus (TMN), which express GluR1, GluR2, and GluR4 AMPA receptor subunits (Sergeeva et al 2004) and the serotonin 2C receptor (Sergeeva et al 2003). As GluR1 subunit pre-mRNA does not undergo editing (Lomeli et al 1994), we determined the extent of editing at known sites of mRNAs encoding for the 5-HT 2C R, as well as GluR2 and GluR4 subunits of the AMPA receptor.…”