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Anesthetic diffusion through lipid membranes depends on the protonation rate.

Pérez-Isidoro R, Sierra-Valdez FJ, Ruiz-Suárez JC - Sci Rep (2014)

Bottom Line: Indeed, such rate modulates the diffusion speed of anesthetics into lipid membranes; low protonation rates enhance the diffusion for local anesthetics while high ones reduce it.We show also that there is a pH and membrane phase dependence on the local anesthetic diffusion across multiple lipid bilayers.Based on our findings we incorporate a new clue that may advance our understanding of the anesthetic phenomenon.

View Article: PubMed Central - PubMed

Affiliation: CINVESTAV-Monterrey, PIIT, Nuevo León, 66600, México.

ABSTRACT
Hundreds of substances possess anesthetic action. However, despite decades of research and tests, a golden rule is required to reconcile the diverse hypothesis behind anesthesia. What makes an anesthetic to be local or general in the first place? The specific targets on proteins, the solubility in lipids, the diffusivity, potency, action time? Here we show that there could be a new player equally or even more important to disentangle the riddle: the protonation rate. Indeed, such rate modulates the diffusion speed of anesthetics into lipid membranes; low protonation rates enhance the diffusion for local anesthetics while high ones reduce it. We show also that there is a pH and membrane phase dependence on the local anesthetic diffusion across multiple lipid bilayers. Based on our findings we incorporate a new clue that may advance our understanding of the anesthetic phenomenon.

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Related in: MedlinePlus

Diffusion kinetics and membrane-phase dependence diffusion of TCC.(a) A sequence of 65 calorimetric profiles of the TCC diffusion kinetics in both ‘free’ (upper curves) and ‘clinical conditions’ (lower curves). The ‘double-phase transition’ is splitted in two sections (H1 and H2) from the midpoint between the two transitions. The time between scans was about 36 min. (b) Enthalpies of H1 and H2 as function of time for both ‘free’ (black circles) and ‘clinical conditions’ (blue triangles). Best-fit models are indicated respectively, from where the diffusion coefficient, κ, for ‘clinical’ (2.35) and ‘free conditions’ (0.45) was obtained. Total calorimetric enthalpy values (ΔHmax) were ~36.7 and ~34.2 kJ/mol for ‘clinical’ (dashed line) and ‘free conditions’ (dotted line), respectively. Error bars show the standard deviation. (c) Enthalpies of H1 and H2 as function of time for experiments performed in gel phase (25°C; black circles), fluid phase (41.8°C; red triangles) and phase-transition temperature (55°C; blue squares), at ‘free conditions’. The κ values for phase transition temperature (9.5), fluid phase (1.7) and gel phase (0.28) were obtained from the diffusion model fit. Only four representative stages from the complete kinetics were carried out to describe the membrane-phase dependence in the three respective conditions. TCC was added to MLV and twelve independent experiments were incubated the required time at their respective temperature. TCC was always used at 25 mM and the solution was adjusted to approximately pH 5 (HCl/NaOH).
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f3: Diffusion kinetics and membrane-phase dependence diffusion of TCC.(a) A sequence of 65 calorimetric profiles of the TCC diffusion kinetics in both ‘free’ (upper curves) and ‘clinical conditions’ (lower curves). The ‘double-phase transition’ is splitted in two sections (H1 and H2) from the midpoint between the two transitions. The time between scans was about 36 min. (b) Enthalpies of H1 and H2 as function of time for both ‘free’ (black circles) and ‘clinical conditions’ (blue triangles). Best-fit models are indicated respectively, from where the diffusion coefficient, κ, for ‘clinical’ (2.35) and ‘free conditions’ (0.45) was obtained. Total calorimetric enthalpy values (ΔHmax) were ~36.7 and ~34.2 kJ/mol for ‘clinical’ (dashed line) and ‘free conditions’ (dotted line), respectively. Error bars show the standard deviation. (c) Enthalpies of H1 and H2 as function of time for experiments performed in gel phase (25°C; black circles), fluid phase (41.8°C; red triangles) and phase-transition temperature (55°C; blue squares), at ‘free conditions’. The κ values for phase transition temperature (9.5), fluid phase (1.7) and gel phase (0.28) were obtained from the diffusion model fit. Only four representative stages from the complete kinetics were carried out to describe the membrane-phase dependence in the three respective conditions. TCC was added to MLV and twelve independent experiments were incubated the required time at their respective temperature. TCC was always used at 25 mM and the solution was adjusted to approximately pH 5 (HCl/NaOH).

Mentions: To evaluate the time diffusion of local anesthetics through multiple bilayers, TCC was selected for this analysis due to its large ΔTm at clinical concentrations (25 mM). A set of 65 successive heating DSC scans of TCC in MLV was obtained to analyze the diffusive kinetics. The whole diffusion kinetics in ‘clinical conditions’ is compared with a ‘free conditions’ case (free from NaCl and Phe) (Fig. 3a). From the ‘double-phase transition’, the left peak (H1) corresponds to TCC-perturbed membranes and the right peak (H2) to pure membranes not yet doped by TCC. It is clearly observed that TCC considerably induces more membrane disorder in ‘clinical’ than in ‘free conditions’. However, regardless the ΔTm induced by TCC, both kinetic profiles evidence that while H1 increases, H2 decreases. After a very long time, the homogeneous distribution of TCC along the multiple bilayers leads to the disappearance of the H2 peak. This final state is equivalent to both the LUV-TCC case (Fig. 1c) or if TCC is added to MLV from the hydration process (Supplementary Fig. S1). Calorimetric enthalpy (ΔH), area under the curve, was calculated for each DSC scan. The total ΔH was separated in two sections from the midpoint between the two transitions, where the left peak area corresponds to H1 and the right peak area to H2 (Fig. 3b). Then, individual calorimetric enthalpies were monitored with time. The total ΔH (ΔHmax) was always conserved: ΔHmax = ΔH1 + ΔH2; where ΔHmax is approximately 36.7 and 34.2 kJ/mol for ‘clinical’ and ‘free conditions’, respectively. Heuristically, in the case of the H1 peak, a diffusion model to best fit our experimental results is: where κ is a parameter related to how fast the drug penetrates into the bilayers (for simplicity, we call it diffusion coefficient). On the other hand, from the conservation of ΔHmax, it is easy to note that: From equations (1) and (2), the best-fitted values for κ, at ‘clinical’ and ‘free conditions’, were 2.35 and 0.45, respectively (Fig. 3b). These results therefore suggest that ‘clinical conditions’ induce a faster diffusion.


Anesthetic diffusion through lipid membranes depends on the protonation rate.

Pérez-Isidoro R, Sierra-Valdez FJ, Ruiz-Suárez JC - Sci Rep (2014)

Diffusion kinetics and membrane-phase dependence diffusion of TCC.(a) A sequence of 65 calorimetric profiles of the TCC diffusion kinetics in both ‘free’ (upper curves) and ‘clinical conditions’ (lower curves). The ‘double-phase transition’ is splitted in two sections (H1 and H2) from the midpoint between the two transitions. The time between scans was about 36 min. (b) Enthalpies of H1 and H2 as function of time for both ‘free’ (black circles) and ‘clinical conditions’ (blue triangles). Best-fit models are indicated respectively, from where the diffusion coefficient, κ, for ‘clinical’ (2.35) and ‘free conditions’ (0.45) was obtained. Total calorimetric enthalpy values (ΔHmax) were ~36.7 and ~34.2 kJ/mol for ‘clinical’ (dashed line) and ‘free conditions’ (dotted line), respectively. Error bars show the standard deviation. (c) Enthalpies of H1 and H2 as function of time for experiments performed in gel phase (25°C; black circles), fluid phase (41.8°C; red triangles) and phase-transition temperature (55°C; blue squares), at ‘free conditions’. The κ values for phase transition temperature (9.5), fluid phase (1.7) and gel phase (0.28) were obtained from the diffusion model fit. Only four representative stages from the complete kinetics were carried out to describe the membrane-phase dependence in the three respective conditions. TCC was added to MLV and twelve independent experiments were incubated the required time at their respective temperature. TCC was always used at 25 mM and the solution was adjusted to approximately pH 5 (HCl/NaOH).
© Copyright Policy - open-access
Related In: Results  -  Collection

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f3: Diffusion kinetics and membrane-phase dependence diffusion of TCC.(a) A sequence of 65 calorimetric profiles of the TCC diffusion kinetics in both ‘free’ (upper curves) and ‘clinical conditions’ (lower curves). The ‘double-phase transition’ is splitted in two sections (H1 and H2) from the midpoint between the two transitions. The time between scans was about 36 min. (b) Enthalpies of H1 and H2 as function of time for both ‘free’ (black circles) and ‘clinical conditions’ (blue triangles). Best-fit models are indicated respectively, from where the diffusion coefficient, κ, for ‘clinical’ (2.35) and ‘free conditions’ (0.45) was obtained. Total calorimetric enthalpy values (ΔHmax) were ~36.7 and ~34.2 kJ/mol for ‘clinical’ (dashed line) and ‘free conditions’ (dotted line), respectively. Error bars show the standard deviation. (c) Enthalpies of H1 and H2 as function of time for experiments performed in gel phase (25°C; black circles), fluid phase (41.8°C; red triangles) and phase-transition temperature (55°C; blue squares), at ‘free conditions’. The κ values for phase transition temperature (9.5), fluid phase (1.7) and gel phase (0.28) were obtained from the diffusion model fit. Only four representative stages from the complete kinetics were carried out to describe the membrane-phase dependence in the three respective conditions. TCC was added to MLV and twelve independent experiments were incubated the required time at their respective temperature. TCC was always used at 25 mM and the solution was adjusted to approximately pH 5 (HCl/NaOH).
Mentions: To evaluate the time diffusion of local anesthetics through multiple bilayers, TCC was selected for this analysis due to its large ΔTm at clinical concentrations (25 mM). A set of 65 successive heating DSC scans of TCC in MLV was obtained to analyze the diffusive kinetics. The whole diffusion kinetics in ‘clinical conditions’ is compared with a ‘free conditions’ case (free from NaCl and Phe) (Fig. 3a). From the ‘double-phase transition’, the left peak (H1) corresponds to TCC-perturbed membranes and the right peak (H2) to pure membranes not yet doped by TCC. It is clearly observed that TCC considerably induces more membrane disorder in ‘clinical’ than in ‘free conditions’. However, regardless the ΔTm induced by TCC, both kinetic profiles evidence that while H1 increases, H2 decreases. After a very long time, the homogeneous distribution of TCC along the multiple bilayers leads to the disappearance of the H2 peak. This final state is equivalent to both the LUV-TCC case (Fig. 1c) or if TCC is added to MLV from the hydration process (Supplementary Fig. S1). Calorimetric enthalpy (ΔH), area under the curve, was calculated for each DSC scan. The total ΔH was separated in two sections from the midpoint between the two transitions, where the left peak area corresponds to H1 and the right peak area to H2 (Fig. 3b). Then, individual calorimetric enthalpies were monitored with time. The total ΔH (ΔHmax) was always conserved: ΔHmax = ΔH1 + ΔH2; where ΔHmax is approximately 36.7 and 34.2 kJ/mol for ‘clinical’ and ‘free conditions’, respectively. Heuristically, in the case of the H1 peak, a diffusion model to best fit our experimental results is: where κ is a parameter related to how fast the drug penetrates into the bilayers (for simplicity, we call it diffusion coefficient). On the other hand, from the conservation of ΔHmax, it is easy to note that: From equations (1) and (2), the best-fitted values for κ, at ‘clinical’ and ‘free conditions’, were 2.35 and 0.45, respectively (Fig. 3b). These results therefore suggest that ‘clinical conditions’ induce a faster diffusion.

Bottom Line: Indeed, such rate modulates the diffusion speed of anesthetics into lipid membranes; low protonation rates enhance the diffusion for local anesthetics while high ones reduce it.We show also that there is a pH and membrane phase dependence on the local anesthetic diffusion across multiple lipid bilayers.Based on our findings we incorporate a new clue that may advance our understanding of the anesthetic phenomenon.

View Article: PubMed Central - PubMed

Affiliation: CINVESTAV-Monterrey, PIIT, Nuevo León, 66600, México.

ABSTRACT
Hundreds of substances possess anesthetic action. However, despite decades of research and tests, a golden rule is required to reconcile the diverse hypothesis behind anesthesia. What makes an anesthetic to be local or general in the first place? The specific targets on proteins, the solubility in lipids, the diffusivity, potency, action time? Here we show that there could be a new player equally or even more important to disentangle the riddle: the protonation rate. Indeed, such rate modulates the diffusion speed of anesthetics into lipid membranes; low protonation rates enhance the diffusion for local anesthetics while high ones reduce it. We show also that there is a pH and membrane phase dependence on the local anesthetic diffusion across multiple lipid bilayers. Based on our findings we incorporate a new clue that may advance our understanding of the anesthetic phenomenon.

Show MeSH
Related in: MedlinePlus