Brugada syndrome (BrS) is a condition defined by ST-segment alteration in right precordial leads and a risk of sudden death. Because BrS is often associated with right bundle branch block and the TRPM4 gene is involved in conduction blocks, we screened TRPM4 for anomalies in BrS cases. The DNA of 248 BrS cases with no SCN5A mutations were screened for TRPM4 mutations. Among this cohort, 20 patients had 11 TRPM4 mutations. Two mutations were previously associated with cardiac conduction blocks and 9 were new mutations (5 absent from ∼14′000 control alleles and 4 statistically more prevalent in this BrS cohort than in control alleles). In addition to Brugada, three patients had a bifascicular block and 2 had a complete right bundle branch block. Functional and biochemical studies of 4 selected mutants revealed that these mutations resulted in either a decreased expression (p.Pro779Arg and p.Lys914X) or an increased expression (p.Thr873Ile and p.Leu1075Pro) of TRPM4 channel. TRPM4 mutations account for about 6% of BrS. Consequences of these mutations are diverse on channel electrophysiological and cellular expression. Because of its effect on the resting membrane potential, reduction or increase of TRPM4 channel function may both reduce the availability of sodium channel and thus lead to BrS.

Abstract-We have characterized modulation of I Ca by Ca 2ϩ at the t-tubules (ie, in control cells) and surface sarcolemma (ie, in detubulated cells) of cardiac ventricular myocytes, using the whole-cell patch clamp technique to record I Ca . I Ca inactivation was significantly slower in detubulated cells than in control cells (27.1Ϯ7.8 ms, nϭ22, versus 16.4Ϯ7.9 ms, nϭ22; PϽ0.05). In atrial myocytes, which lack t-tubules, I Ca inactivation was not changed by the treatment used to produce detubulation. In the presence of ryanodine or BAPTA, or when Ba 2ϩ was used as the charge carrier, the rate of inactivation was not significantly different in control and detubulated cells. Frequency-dependent facilitation occurred in control cells but not in detubulated cells, and was abolished by ryanodine. These results suggest that Ca 2ϩ released from the SR has a greater effect on I Ca in the t-tubules than at the surface sarcolemma. This does not appear to be due to differences in local Ca 2ϩ release from the SR, because the gain of Ca 2ϩ release was not significantly different in control and detubulated cells. These data suggest that the t-tubules are a key site for the regulation of transsarcolemmal releases. 2 The transverse (t-) tubules of ventricular myocytes play an important role in this process. These tubules are invaginations of the sarcolemma that occur at the Z-line, perpendicular to the longitudinal axis of the cell (see review). 3 Functional and immunohistochemical data suggest that I Ca occurs predominantly in the t-tubules, adjacent to RyRs, which are also located predominantly at the t-tubules. 4 -6 Thus, it appears that the t-tubules are the major site of Ca 2ϩ entry and hence Ca 2ϩ release in cardiac ventricular myocytes. Conversely, Ca 2ϩ released by the SR can modulate I Ca ; this plays an important role in cellular Ca 2ϩ homeostasis, controlling Ca 2ϩ entry via negative feedback. 7,8 However, it is unknown whether the efficacy of coupling between SR Ca 2ϩ release and I Ca is the same in the t-tubule and surface membranes, so that the relative importance of these sites in cellular Ca 2ϩ homeostasis is unknown. We have, therefore, investigated the regulation of I Ca by Ca 2ϩ released from the SR in normal ventricular myocytes, in which I Ca triggers Ca 2ϩ release predominantly at the t-tubules, and in myocytes in which the t-tubules have been physically and functionally uncoupled from the surface membrane (detubulated), 5 in which Ca 2ϩ release occurs predominantly at the surface membrane. 9

Hypoxia and re-oxygenation-induced EADs can be generated in the mouse heart model. 9-Phenanthrol abolished EADs, which strongly suggests the involvement of TRPM4 in the generation of EAD. This identifies non-selective cation channels inhibitors as new pharmacological candidates in the treatment of arrhythmias.

No abstract

TRPM4 forms a non-selective cation channel activated by internal Ca(2+). Its functional expression was demonstrated in cardiomyocytes of several mammalian species including humans, but the channel is also present in many other tissues. The recent characterization of the TRPM4 inhibitor 9-phenanthrol, and the availability of transgenic mice have helped to clarify the role of TRPM4 in cardiac electrical activity, including diastolic depolarization from the sino-atrial node cells in mouse, rat, and rabbit, as well as action potential duration in mouse cardiomyocytes. In rat and mouse, pharmacological inhibition of TRPM4 prevents cardiac ischaemia-reperfusion injuries and decreases the occurrence of arrhythmias. Several studies have identified TRPM4 mutations in patients with inherited cardiac diseases including conduction blocks and Brugada syndrome. This review identifies TRPM4 as a significant actor in cardiac electrophysiology.

Transient Receptor Potential (TRP) proteins are non-selective cationic channels with a consistent Ca(2+)-permeability, except for TRPM4 and TRPM5 that are not permeable to this ion. However, Ca(2+) is a major regulator of their activity since both channels are activated by a rise in internal Ca(2+). Thus TRPM4 and TRPM5 are responsible for most of the Ca(2+)-activated non-selective cationic currents (NSC(Ca)) recorded in a large variety of tissues. Their activation induces cell-membrane depolarization that modifies the driving force for ions as well as activity of voltage gated channels and thereby strongly impacts cell physiology. In the last few years, the ubiquitously expressed TRPM4 channel has been implicated in insulin secretion, the immune response, constriction of cerebral arteries, the activity of inspiratory neurons and cardiac dysfunction. Conversely, TRPM5 whose expression is more restricted, has until now been mainly implicated in taste transduction.

The action potential of cardiac ventricular myocytes is characterized by its long duration, mainly due to Ca flux through L-type Ca channels. Ca entry also serves to trigger the release of Ca from the sarcoplasmic reticulum. The aim of this study was to investigate the role of cell membrane invaginations called transverse (T)-tubules in determining Ca influx and action potential duration in cardiac ventricular myocytes. We used the whole cell patch clamp technique to record electrophysiological activity in intact rat ventricular myocytes (i.e., from the T-tubules and surface sarcolemma) and in detubulated myocytes (i.e., from the surface sarcolemma only). Action potentials were significantly shorter in detubulated cells than in control cells. In contrast, resting membrane potential and action potential amplitude were similar in control and detubulated myocytes. Experiments under voltage clamp using action potential waveforms were used to quantify Ca entry via the Ca current. Ca entry after detubulation was reduced by approximately 60%, a value similar to the decrease in action potential duration. We calculated that Ca influx at the T-tubules is 1.3 times that at the cell surface (4.9 vs. 3.8 micromol/L cytosol, respectively) during a square voltage clamp pulse. In contrast, during a cardiac action potential, Ca entry at the T-tubules is 2.2 times that at the cell surface (3.0 vs. 1.4 micromol/L cytosol, respectively). However, more Ca entry occurs per microm(2) of junctional membrane at the cell surface than in the T-tubules (in nM/microm(2): 1.43 vs. 1.06 during a cardiac action potential). This difference is unlikely to be due to a difference in the number of Ca channels/junction at each site because we estimate that the same number of Ca channels is present at cell surface and T-tubule junctions ( approximately 35). This study provides the first evidence that the T-tubules are a key site for the regulation of action potential duration in ventricular cardiac myocytes. Our data also provide the first direct measurements of T-tubular Ca influx, which are consistent with the idea that cardiac excitation-contraction coupling largely occurs at the T-tubule dyadic clefts.

The 52‐kDa SSA/Ro (Ro52) ribonucleoprotein is an antigenic target strongly associated with the autoimmune response in mothers whose children develop neonatal lupus and congenital heart block. When sera from patients with systemic lupus erythematosus were used as autoimmune controls in an enzyme immunoassay to screen for antibodies against the human serotoninergic 5‐HT4‐receptor, a high correlation was found between the presence of anti‐Ro52 protein antibodies in such sera and antibodies reacting with a synthetic peptide, corresponding to the second extracellular loop of the human 5‐HT4 receptor (amino acid residues 165–185). Homology scanning between the 5‐HT4 peptide and the sequence of the Ro52 protein indicated two potential common epitopes located between residues 365 and 396 of the Ro52 protein. Cross‐reactivity was found between the peptide derived from the 5‐HT4 receptor, and a peptide corresponding to residues 365–382 of the Ro52 protein. Autoantibodies, affinity‐purified on the 5‐HT4 receptor peptide, specifically recognized both the Ro52 protein and the 5‐HT4 receptor protein in immunoblots. The affinity‐purified antibodies antagonized the serotonin‐induced L‐type Ca channel activation on human atrial cells. This effect could explain the electrophysiological abnormalities in neonatal lupus.