M2 receptor gene sequencing After sequencing the selleck chem Nilotinib cholinergic receptor M2 gene, we detected a single nucleotide mutation from thymine to guanine (T��G) in position 1311 (Fig. 6a). This mutation transformed the normal CCT codon into CCG, both encoding for the proline amino-acid. The two codons were not equally distributed among animals. Over the 46 rabbits tested, 17 had the wild genotype CCT/CCT, 12 were heterozygous CCT/CCG and 17 were homozygous CCG/CCG. None of the 11 normal rabbits had the mutation, while it was present in 29/35, i.e., 83% of the vagal hyperreactive rabbits (Fig. 6b). The mean R-R interval in normal rabbits was 3318��1336 ms; mean R-R intervals in hyperreactive rabbits were 24700��6855 ms (wild genotype CCT/CCT), 16650��6572 ms (heterozygous CCT/CCG) and 21096��7759 ms (homozygous CCG/CCG) (Fig.

6b). Figure 6 Polymorphism of the M2 cholinergic muscarinic receptor gene of normal (N) and vagal hyperreactive (H) rabbits. Discussion In a previous study [12], we showed that spontaneous vagal pauses were observed in a particular strain of adult rabbits (12�C14 weeks of age) which can therefore be used as an experimental model of vagal hyperreactivity. In the experimental conditions of the present study, the PNE test was used to screen hyperreactive animals. To prevent sudden death due to arrhythmogenic complications of PNE, the animals were treated with the ��-blocking drug, propranolol. A unique bolus dose of the latter was delivered shortly before the PNE test. As shown previously, maximal R-R interval is a reliable index to assess the vagal reactivity in these animals [12].

In binding experiments, we showed that the densities of both M2 and M3 muscarinic receptors were enhanced in the heart of rabbits displaying exacerbated vagal responses. A similar increase of M2 mRNA was observed in peripheral mononuclear white blood cells. As the sequence of the M3 receptor gene was not known, PCR experiments regarding mRNA expression could not be performed. A significant correlation was established between the severity of the bradycardia and the cardiac muscarinic receptor expression level indicating that vagal hyperreactivity was highly dependent on muscarinic receptor density. The increase in the M2 subtype is in line with the well-established negative chronotropic effects of these receptors. We also found a significant increase in M3 receptor expression.

M3 receptors have been identified Anacetrapib in hearts of several mammalian species, including humans [16], [20]�C[22], and recent studies showed that M3 stimulation mediates K+ currents in cardiac cells [23], [24]. Interestingly, no overlap between the muscarinic receptor expression in normal and diseased animals was observed and muscarinic receptor overexpression was detected in all hyperreactive animals, suggesting that it is a primary cardiac abnormality underlying vagal hyperreactivity.