Korean J Physiol Pharmacol.  2025 May;29(3):373-384. 10.4196/kjpp.24.363.

Quetiapine competitively inhibits 5-HT₃ receptor-mediated currents in NCB20 neuroblastoma cells

• Park YS
• Kim GM
• Sung HJ
• Yu JY
• Sung KW

• KMID: 2567194
• DOI: http://doi.org/10.4196/kjpp.24.363

Abstract

The 5-hydroxytryptamine type₃ (5-HT₃ ) receptor, a ligand-gated ion channel, plays a critical role in synaptic transmission. It has been implicated in various neuropsychiatric disorders. This study aimed to elucidate the mechanism by which quetiapine, an atypical antipsychotic, could inhibit 5-HT₃ receptor-mediated currents in NCB20 neuroblastoma cells. Whole-cell patch-clamp recordings were used to study effects of quetiapine on receptor ion channel kinetics and its competitive antagonism. Co-application of quetiapine shifted 5-HT concentration-response curve rightward, significantly increasing the EC₅₀ without altering the maximal response (E_max ), suggesting a competitive inhibition. Quetiapine's IC₅₀ varied with 5-HT concentration and treatment condition. The IC₅₀ value of quetiapine was 0.58 μM with 3 μM 5-HT and 25.23 μM with 10 μM 5-HT, indicating an inverse relationship between quetiapine efficacy and agonist concentration. Pretreatment of quetiapine significantly enhanced its inhibitory potency, reducing its IC₅₀ from 25.23 μM to 0.20 μM. Interaction kinetics experiments revealed an IC₅₀ of 5.17 μM for an open state of the 5-HT₃ receptor, suggesting weaker affinity during receptor activation. Quetiapine also accelerated receptor deactivation and desensitization, suggesting that it could stabilize the receptor in non-conducting states. Additionally, quetiapine significantly prolonged recovery from desensitization without affecting recovery from deactivation, demonstrating its selective impact on receptor kinetics. Inhibition of the 5-HT₃ receptor by quetiapine was voltage-independent, and quetiapine exhibited no usedependency, further supporting its role as a competitive antagonist. These findings provide insights into inhibitory mechanism of quetiapine on 5-HT₃ receptor and suggest its potential therapeutic implications for modulating serotonergic pathways in neuropsychiatric disorders.

Keyword

Competitive binding; Patch-clamp techniques; Quetiapine fumarate; Schizophrenia; 5-HT₃ receptor

Figure

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  Fig. 1 Effect of quetiapine on effective concentration 50% (EC50) and maximal effect (Emax) of 5-HT (5-hydroxytryptamine)-induced 5-HT3 receptor-mediated currents. This figure shows representative whole-cell voltage-clamp recordings of 5-HT3 receptor-mediated currents in NCB20 neuroblastoma cells, illustrating effects of quetiapine co-applied with various concentrations of 5-HT. Increasing concentrations of 5-HT (0.3, 1, 3, 10, 30, and 100 μM) were applied alone (left traces in A, B, C) or co-applied with quetiapine (right traces in A, B, C). (A) 5-HT3 receptor-mediated currents evoked by 5-HT alone (left) or co-applied with 3 μM quetiapine (right). Quetiapine significantly reduced the amplitude of currents evoked by low concentrations (0.3, 1, and 3 μM) of 5-HT, while the peak current amplitude induced by higher concentrations (10, 30, and 100 μM) of 5-HT exhibited only a minimal reduction. (B, C) 5-HT3 receptor-mediated currents evoked by 5-HT alone (left) or co-applied with 10 or 30 μM quetiapine (right). A similar reduction in current amplitude was observed at lower 5-HT concentrations (0.3, 1, and 3 μM), with a less pronounced effect at higher 5-HT concentrations (10, 30, and 100 μM). (D) Concentration-response curves for 5-HT alone or co-applied with quetiapine (3, 10, and 30 μM). Quetiapine shifted the concentration-response curve of 5-HT to the right in a concentration-dependent manner, indicating a decrease in 5-HT potency. EC50 values increased significantly with quetiapine co-application, from 2.64 ± 0.06 μM for 5-HT alone (n = 26) to 4.70 ± 0.23 μM in the presence of 3 μM quetiapine (n = 10), 6.61 ± 0.20 μM in the presence of 10 μM quetiapine (n = 10), and 9.99 ± 1.27 μM in the presence of 30 μM quetiapine (n = 9). Emax wase unaffected by quetiapine, suggesting that quetiapine could modulate receptor sensitivity without reducing the maximum response. Data are presented as mean ± SEM. Statistical significance was determined using one-way ANOVA (F(3,51) = 48.95, p < 0.001).

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  Fig. 2 Concentration-dependent inhibition of 5-hydroxytryptamine type3 (5-HT3) receptor-mediated currents by quetiapine. (A) Representative 5-HT3 receptor currents evoked by 3 μM 5-HT alone (gray traces) or co-applied with quetiapine at concentrations ranging from 0.1 to 30 μM (black traces). Quetiapine co-application resulted in a concentration-dependent reduction in the peak amplitude of 5-HT3 receptor-mediated currents, with a more prominent inhibition at higher concentrations of quetiapine. (B) Representative 5-HT3 receptor currents evoked by 10 μM 5-HT alone (gray traces) or co-applied with quetiapine at concentrations ranging from 0.3 to 100 μM (black traces). As in panel A, quetiapine effectively reduced current amplitude in a concentration-dependent manner, although the inhibition was less profound compared to 3 μM 5-HT. (C) Representative traces showing 5-HT3 receptor currents evoked by 10 μM 5-HT (gray traces) or pretreated with quetiapine for 5 sec before co-application with 10 μM 5-HT (black traces). Pretreatment with quetiapine enhanced the inhibitory effect, resulting in a more substantial reduction in peak current at lower quetiapine concentrations. This suggests that quetiapine can preferentially bind to closed states of the 5-HT3 receptor ion channel, thereby enhancing its inhibitory effect when pre-applied. (D) Concentration-response curves summarizing quetiapine’s inhibitory effects under three conditions: co-application with 3 μM 5-HT (gray circles), co-application with 10 μM 5-HT (open circles), and pretreatment of quetiapine for 5 sec before co-application with 10 μM 5-HT (filled circles). IC50 values was significantly lower with 3 μM 5-HT (0.58 ± 0.08 μM, n = 10) compared to that with 10 μM 5-HT (25.23 ± 3.28 μM, n = 8) (p < 0.001). Pretreatment with quetiapine further reduced the IC50 to 0.20 ± 0.01 μM (n = 10), indicating an enhanced inhibitory effect (unpaired t-test, p < 0.001). The Hill slope remained consistent across conditions, suggesting that quetiapine could modulate receptor sensitivity without affecting the cooperativity of 5-HT binding. Data are presented as mean ± SEM.

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  Fig. 3 Effects of quetiapine on deactivation and desensitization of 5-hydroxytryptamine type3 (5-HT3) receptor-mediated currents. (A) Representative superimposed traces of 5-HT3 receptor-mediated currents evoked by a 15 msec application of 10 μM 5-HT alone (gray trace) or co-applied with quetiapine at increasing concentrations (1, 3, 10, and 30 μM; black traces). Quetiapine concentration-dependently accelerated the deactivation of 5-HT3 receptor-mediated currents. (B) Representative superimposed traces of 5-HT3 receptor-mediated currents evoked by a 20 sec application of 10 μM 5-HT alone (gray trace) or co-applied with quetiapine at increasing concentrations (1, 3, 10, and 30 μM; black traces). Quetiapine accelerated the desensitization of 5-HT3 receptor-mediated currents, with faster decay observed at higher concentrations. (C) Summarized data for decay time constant (τ) of deactivation and desensitization plotted against quetiapine concentration. For deactivation, the decay τ for 5-HT alone was 1,991 ± 207.4 ms, which decreased to 1,752 ± 190.3 ms in the presence of 1 μM quetiapine, 1,612 ± 170.4 ms in the presence of 3 μM quetiapine, 1,467 ± 164.5 ms in the presence of 10 μM quetiapine, and 1,303 ± 139.7 ms in the presence of 30 μM quetiapine (open circle). For desensitization, the τ for 5-HT alone was 3,865 ± 511.5 ms, which decreased to 2,247 ± 316.6 ms in the presence of 1 μM quetiapine, 1,312 ± 157.7 ms in the presence of 3 μM quetiapine, 507.9 ± 48.02 ms in the presence of 10 μM quetiapine, and 214.1 ± 20.44 ms in the presence of 30 μM quetiapine (closed circle). Repeated measures one-way ANOVA showed a significant effect of quetiapine on both deactivation (F(1.253, 10.02) = 47.86, p < 0.001, n = 9) and desensitization (F(1.019, 8.152) = 51.23, p < 0.001, n = 9). Data are presented as mean ± SEM. *** indicates p < 0.001 as determined by Dunnett’s multiple comparisons test, comparing each quetiapine concentration with 5-HT alone.

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  Fig. 4 Effects of quetiapine on recovery from deactivation and desensitization of 5-hydroxytryptamine type3 (5-HT3) receptor-mediated currents. (A) Representative superimposed traces of 5-HT3 receptor-mediated currents showing recovery from deactivation. The first pulse was a brief 20 msec application of 10 μM 5-HT (gray arrows), followed by a 2.5 sec pulse (black arrows) after varying inter-pulse intervals (1, 5, 10, 30, and 60 sec). The left panel shows currents evoked by 10 μM 5-HT alone, while the right panel shows currents with co-application of 10 μM quetiapine. (B) Averaged paired pulse ratio (PPR; second peak amplitude/first peak amplitude) plotted against the inter-pulse interval. PPR data were fitted to a one-phase association equation to determine time constants (τ). The τ for 5-HT alone (open circles) was 5.53 ± 0.28 sec (n = 10) and 5.43 ± 0.39 sec for co-application with quetiapine (filled circles, n = 9). Statistical analysis showed no significant difference between the two conditions (p = 0.8437, unpaired t-test), indicating that quetiapine did not alter the recovery from deactivation. (C) Representative superimposed traces of 5-HT3 receptor-mediated currents showing recovery from desensitization. The first pulse was a 5 sec application of 10 μM 5-HT (gray arrows), followed by a 2.5 sec second pulse (black arrows) after varying inter-pulse intervals (1, 5, 10, 30, and 60 sec). The left panel shows currents evoked by 10 μM 5-HT alone and the right panel displays currents evoked by co-application with 10 μM quetiapine. (D) Summarized PPR plotted against the inter-pulse interval, with data fitted to a one-phase association equation. The τ values was 6.88 ± 0.41 sec for 5-HT alone (open circles, n = 9) and 9.85 ± 0.75 sec for co-application with quetiapine (closed circles, n = 9). Statistical analysis revealed a significant difference between the two conditions (p < 0.01, unpaired t-test), indicating that quetiapine prolonged the recovery from desensitization of 5-HT3 receptor-mediated currents. Data are presented as mean ± SEM.

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  Fig. 5 Effect of quetiapine on current-voltage (I-V) relationship of 5-hydroxytryptamine type3 (5-HT3) receptor-mediated currents. (A) Representative current traces recorded at holding potentials of –50, –30, –10, +10, and +30 mV. The left panel shows currents evoked by 10 μM 5-HT alone, while the right panel displays currents evoked by co-application with 10 μM quetiapine. (B) Averaged normalized peak amplitudes of 5-HT3 receptor currents plotted against the holding potential (VHolding). The reversal potential was determined by extrapolating the I-V curve to the point where the current amplitude crossed zero. The reversal potential was 2.38 ± 0.20 mV for 10 μM 5-HT alone (open circles) and 2.51 ± 0.19 mV for co-application with 10 μM quetiapine (closed circles). This difference was not statistically significant (n = 9, p = 0.1617, paired t-test). (C) Peak amplitude ratio, defined as the ratio of the peak current amplitude for 5-HT with quetiapine co-application to that for 5-HT alone, plotted across different holding potentials. Peak amplitude ratios were consistent across all tested holding potentials. Statistical analysis by repeated measures one-way ANOVA revealed no significant effect of holding potential on the peak amplitude ratio (F(2.046, 16.37) = 1.045, p = 0.3756). Data are presented as mean ± SEM.

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  Fig. 6 Association kinetics of quetiapine with open states of 5-hydroxytryptamine type3 (5-HT3) receptor. (A) Superimposed current traces recorded during a protocol in which 3 μM 5-HT was applied for 16 sec, and quetiapine at varying concentrations (1, 3, 10, 30 μM) was co-applied with 5-HT for 8 sec, starting 3 sec after the onset of 5-HT application. These traces demonstrated that quetiapine accelerated the decay of 5-HT3 receptor currents during co-application in a concentration-dependent manner. Currents reappeared upon discontinuation of quetiapine co-application at concentrations higher than 3 μM. (B) Time constant (τ) of the current decay plotted against quetiapine concentration, showing that the τ decreased with increasing quetiapine concentration (F(1.436,12.92) = 191.7, p < 0.001, repeated measures one-way ANOVA). Decay of the current during quetiapine co-application was fitted to a single exponential function to obtain decay τ and analyzed for each concentration of quetiapine. (C) The reciprocal of the time constant (1/τ) plotted as a function of quetiapine concentration. According to the first-order blocking scheme, the slope of the linear fit represents the association rate constant (k+1), while the y-intercept corresponds to the dissociation rate constant (k-1) for the open 5-HT3 receptor. The linear fit yielded a slope of 0.08776 and a y-intercept of 0.4539. The apparent dissociation constant (Kd) of quetiapine calculated as the ratio of k-1 to k+1 was determined to be 5.17 μM. Data are expressed as mean ± SEM.

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  Fig. 7 Analysis of use-dependence of quetiapine’s inhibitory action on 5-hydroxytryptamine type3 (5-HT3) receptor currents. (A) Representative current traces of 10 μM 5-HT applied in 2 sec pulses at 30 sec intervals for a total of 16 applications. Quetiapine (10 μM) was co-applied during the 4th to 8th applications, as indicated by black bars. Gray traces represent currents evoked by 5-HT alone, while black traces illustrate currents during co-application of quetiapine. (B) Time course of averaged normalized peak amplitudes during the 16 consecutive 5-HT applications. The peak amplitude decreased during quetiapine co-application and recovered after its discontinuation. (C) Comparison of normalized peak amplitudes between the 5th and 8th applications. There was no significant difference between the two (5th application: 0.69 ± 0.01; 8th application: 0.67 ± 0.02; p = 0.1340, n = 8), indicating no use-dependent inhibition by quetiapine. Data are presented as mean ± SEM.

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