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Congenital hyperinsulinism and glucose hypersensitivity in homozygous and heterozygous carriers of Kir6.2 (KCNJ11) mutation V290M mutation: K(ATP) channel inactivation mechanism and clinical management.

Loechner KJ, Akrouh A, Kurata HT, Dionisi-Vici C, Maiorana A, Pizzoferro M, Rufini V, de Ville de Goyet J, Colombo C, Barbetti F, Koster JC, Nichols CG - Diabetes (2010)

Bottom Line: In vitro expression in COSm6 cells supports the mutation resulting in an inactivating phenotype, which leads to significantly reduced activity in intact cells when expressed homomerically, and to a lesser extent when expressed heteromerically with wild-type subunits.The patient continues to be treated successfully with octreotide and amlodipine.V290M results in inactivating K(ATP) channels that underlie HI.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, University of North Carolina School of Medicine, Chapel Hill, USA.

ABSTRACT

Objective: The ATP-sensitive K(+) channel (K(ATP)) controls insulin secretion from the islet. Gain- or loss-of-function mutations in channel subunits underlie human neonatal diabetes and congenital hyperinsulinism (HI), respectively. In this study, we sought to identify the mechanistic basis of K(ATP)-induced HI in two probands and to characterize the clinical course.

Research design and methods: We analyzed HI in two probands and characterized the course of clinical treatment in each, as well as properties of mutant K(ATP) channels expressed in COSm6 cells using Rb efflux and patch-clamp methods.

Results: We identified mutation V290M in the pore-forming Kir6.2 subunit in each proband. In vitro expression in COSm6 cells supports the mutation resulting in an inactivating phenotype, which leads to significantly reduced activity in intact cells when expressed homomerically, and to a lesser extent when expressed heteromerically with wild-type subunits. In one heterozygous proband, a fluoro-DOPA scan revealed a causal focal lesion, indicating uniparental disomy with loss of heterozygosity. In a second family, the proband, homozygous for the mutation, was diagnosed with severe diazoxide-unresponsive hypersinsulinism at 2 weeks of age. The patient continues to be treated successfully with octreotide and amlodipine. The parents and a male sibling are heterozygous carriers without overt clinical HI. Interestingly, both the mother and the sibling exhibit evidence of abnormally enhanced glucose tolerance.

Conclusions: V290M results in inactivating K(ATP) channels that underlie HI. Homozygous individuals may be managed medically, without pancreatectomy. Heterozygous carriers also show evidence of enhanced glucose sensitivity, consistent with incomplete loss of K(ATP) channel activity.

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Mutant V290M channels have unaltered ATP-sensitivity and intact MgADP activation. A: Representative currents recorded by inside-out excised patch-clamp technique from COSm6 cells expressing WT or mutant V290M KATP channels at −50 mV membrane potential (+50 mV pipette). Patches were exposed to different concentrations of ATP as indicated. B: Mean steady-state patch current following isolation (± SEM). C: Steady-state dependence of membrane current on ATP relative to current in the absence of ATP. Solid line represents mutant V290M channels while dashed line represents WT channels fitted with the Hill equation by least-squares method. D: Representative currents recorded by inside-out excised patch-clamp technique from COSm6 cells expressing WT or mutant V290M KATP channels at −50 mV membrane potential (+50 mV pipette). Patches were exposed to different concentrations of ATP, and ADP (in the presence of 0.5. mM free Mg2+) as indicated.
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Figure 4: Mutant V290M channels have unaltered ATP-sensitivity and intact MgADP activation. A: Representative currents recorded by inside-out excised patch-clamp technique from COSm6 cells expressing WT or mutant V290M KATP channels at −50 mV membrane potential (+50 mV pipette). Patches were exposed to different concentrations of ATP as indicated. B: Mean steady-state patch current following isolation (± SEM). C: Steady-state dependence of membrane current on ATP relative to current in the absence of ATP. Solid line represents mutant V290M channels while dashed line represents WT channels fitted with the Hill equation by least-squares method. D: Representative currents recorded by inside-out excised patch-clamp technique from COSm6 cells expressing WT or mutant V290M KATP channels at −50 mV membrane potential (+50 mV pipette). Patches were exposed to different concentrations of ATP, and ADP (in the presence of 0.5. mM free Mg2+) as indicated.

Mentions: The activity of WT and mutant KATP channels was further examined in inside-out membrane patch-clamp experiments (Figs. 3–5). Upon membrane excision, WT channels typically open to a steady-state level with open probability of 0.3–0.5 (20). In marked contrast, V290M channels open but then exhibit a rapid decay in macroscopic current, typically resulting in much smaller steady-state currents than is observed in WT channels. Following patch excision, V290M channels inactivated with time constant of 2.0 ± 0.5 s (n = 5), while WT channels showed essentially no inactivation following excision (Fig. 3B). Steady-state ATP sensitivity of V290M channels was similar to WT (Fig. 4A and C), but current density was considerably lower than WT (Fig. 4B), reflecting the dramatic inactivation that occurs. Physiologically, the major determinant of channel activation is stimulation by Mg-nucleotides and, as shown in Fig. 4D, MgADP activation is intact in V290M channels, although inactivation follows the MgADP activation, reducing steady-state currents in MgADP.


Congenital hyperinsulinism and glucose hypersensitivity in homozygous and heterozygous carriers of Kir6.2 (KCNJ11) mutation V290M mutation: K(ATP) channel inactivation mechanism and clinical management.

Loechner KJ, Akrouh A, Kurata HT, Dionisi-Vici C, Maiorana A, Pizzoferro M, Rufini V, de Ville de Goyet J, Colombo C, Barbetti F, Koster JC, Nichols CG - Diabetes (2010)

Mutant V290M channels have unaltered ATP-sensitivity and intact MgADP activation. A: Representative currents recorded by inside-out excised patch-clamp technique from COSm6 cells expressing WT or mutant V290M KATP channels at −50 mV membrane potential (+50 mV pipette). Patches were exposed to different concentrations of ATP as indicated. B: Mean steady-state patch current following isolation (± SEM). C: Steady-state dependence of membrane current on ATP relative to current in the absence of ATP. Solid line represents mutant V290M channels while dashed line represents WT channels fitted with the Hill equation by least-squares method. D: Representative currents recorded by inside-out excised patch-clamp technique from COSm6 cells expressing WT or mutant V290M KATP channels at −50 mV membrane potential (+50 mV pipette). Patches were exposed to different concentrations of ATP, and ADP (in the presence of 0.5. mM free Mg2+) as indicated.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3012173&req=5

Figure 4: Mutant V290M channels have unaltered ATP-sensitivity and intact MgADP activation. A: Representative currents recorded by inside-out excised patch-clamp technique from COSm6 cells expressing WT or mutant V290M KATP channels at −50 mV membrane potential (+50 mV pipette). Patches were exposed to different concentrations of ATP as indicated. B: Mean steady-state patch current following isolation (± SEM). C: Steady-state dependence of membrane current on ATP relative to current in the absence of ATP. Solid line represents mutant V290M channels while dashed line represents WT channels fitted with the Hill equation by least-squares method. D: Representative currents recorded by inside-out excised patch-clamp technique from COSm6 cells expressing WT or mutant V290M KATP channels at −50 mV membrane potential (+50 mV pipette). Patches were exposed to different concentrations of ATP, and ADP (in the presence of 0.5. mM free Mg2+) as indicated.
Mentions: The activity of WT and mutant KATP channels was further examined in inside-out membrane patch-clamp experiments (Figs. 3–5). Upon membrane excision, WT channels typically open to a steady-state level with open probability of 0.3–0.5 (20). In marked contrast, V290M channels open but then exhibit a rapid decay in macroscopic current, typically resulting in much smaller steady-state currents than is observed in WT channels. Following patch excision, V290M channels inactivated with time constant of 2.0 ± 0.5 s (n = 5), while WT channels showed essentially no inactivation following excision (Fig. 3B). Steady-state ATP sensitivity of V290M channels was similar to WT (Fig. 4A and C), but current density was considerably lower than WT (Fig. 4B), reflecting the dramatic inactivation that occurs. Physiologically, the major determinant of channel activation is stimulation by Mg-nucleotides and, as shown in Fig. 4D, MgADP activation is intact in V290M channels, although inactivation follows the MgADP activation, reducing steady-state currents in MgADP.

Bottom Line: In vitro expression in COSm6 cells supports the mutation resulting in an inactivating phenotype, which leads to significantly reduced activity in intact cells when expressed homomerically, and to a lesser extent when expressed heteromerically with wild-type subunits.The patient continues to be treated successfully with octreotide and amlodipine.V290M results in inactivating K(ATP) channels that underlie HI.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, University of North Carolina School of Medicine, Chapel Hill, USA.

ABSTRACT

Objective: The ATP-sensitive K(+) channel (K(ATP)) controls insulin secretion from the islet. Gain- or loss-of-function mutations in channel subunits underlie human neonatal diabetes and congenital hyperinsulinism (HI), respectively. In this study, we sought to identify the mechanistic basis of K(ATP)-induced HI in two probands and to characterize the clinical course.

Research design and methods: We analyzed HI in two probands and characterized the course of clinical treatment in each, as well as properties of mutant K(ATP) channels expressed in COSm6 cells using Rb efflux and patch-clamp methods.

Results: We identified mutation V290M in the pore-forming Kir6.2 subunit in each proband. In vitro expression in COSm6 cells supports the mutation resulting in an inactivating phenotype, which leads to significantly reduced activity in intact cells when expressed homomerically, and to a lesser extent when expressed heteromerically with wild-type subunits. In one heterozygous proband, a fluoro-DOPA scan revealed a causal focal lesion, indicating uniparental disomy with loss of heterozygosity. In a second family, the proband, homozygous for the mutation, was diagnosed with severe diazoxide-unresponsive hypersinsulinism at 2 weeks of age. The patient continues to be treated successfully with octreotide and amlodipine. The parents and a male sibling are heterozygous carriers without overt clinical HI. Interestingly, both the mother and the sibling exhibit evidence of abnormally enhanced glucose tolerance.

Conclusions: V290M results in inactivating K(ATP) channels that underlie HI. Homozygous individuals may be managed medically, without pancreatectomy. Heterozygous carriers also show evidence of enhanced glucose sensitivity, consistent with incomplete loss of K(ATP) channel activity.

Show MeSH
Related in: MedlinePlus