RNA expression profiling in brains of familial hemiplegic migraine type 1 knock-in mice

Background Various CACNA1A missense mutations cause familial hemiplegic migraine type 1 (FHM1), a rare monogenic subtype of migraine with aura. FHM1 mutation R192Q is associated with pure hemiplegic migraine, whereas the S218L mutation causes hemiplegic migraine, cerebellar ataxia, seizures, and mild head trauma-induced brain edema. Transgenic knock-in (KI) migraine mouse models were generated that carried either the FHM1 R192Q or the S218L mutation and were shown to exhibit increased CaV2.1 channel activity. Here we investigated their cerebellar and caudal cortical transcriptome. Methods Caudal cortical and cerebellar RNA expression profiles from mutant and wild-type mice were studied using microarrays. Respective brain regions were selected based on their relevance to migraine aura and ataxia. Relevant expression changes were further investigated at RNA and protein level by quantitative polymerase chain reaction (qPCR) and/or immunohistochemistry, respectively. Results Expression differences in the cerebellum were most pronounced in S218L mice. Particularly, tyrosine hydroxylase, a marker of delayed cerebellar maturation, appeared strongly upregulated in S218L cerebella. In contrast, only minimal expression differences were observed in the caudal cortex of either mutant mice strain. Conclusion Despite pronounced consequences of migraine gene mutations at the neurobiological level, changes in cortical RNA expression in FHM1 migraine mice compared to wild-type are modest. In contrast, pronounced RNA expression changes are seen in the cerebellum of S218L mice and may explain their cerebellar ataxia phenotype.

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The familial hemiplegic migraine type 1 mutation K1336E affects direct G protein-mediated regulation of neuronal P/Q-type Ca2+ channels

Cephalalgia ◽

10.1177/0333102412475236 ◽

2013 ◽

Vol 33 (6) ◽

pp. 398-407 ◽

Cited By ~ 8

Author(s):

Edgar Garza-López ◽

Ricardo González-Ramírez ◽

María A Gandini ◽

Alejandro Sandoval ◽

Ricardo Felix

Keyword(s):

G Protein ◽

Familial Hemiplegic Migraine ◽

Inactivation Kinetics ◽

Missense Mutations ◽

Hemiplegic Migraine ◽

Gene Encoding ◽

Dominant Form ◽

Hek 293 ◽

Autosomal Dominant Form

Background Familial hemiplegic migraine type 1 (FHM-1) is an autosomal dominant form of migraine with aura characterized by recurrent migraine, hemiparesis and ataxia. FHM-1 has been linked to missense mutations in the CACNA1A gene encoding the pore-forming subunit of the neuronal voltage-gated P/Q-type Ca2+ channel (CaV2.1α1). Methods Here, we explored the effects of the FHM-1 K1336E mutation on G protein-dependent modulation of the recombinant P/Q-type channel. The mutation was introduced into the human CaV2.1α1 subunit and its functional consequences investigated after heterologous expression in HEK-293 cells using patch-clamp recordings. Results Functional analysis of the K1336E mutation revealed a reduction of Ca2+ current densities, a ∼10 mV left-shift in the current-voltage relationship, and the slowing of current inactivation kinetics. When co-expressed along with the human μ-opioid receptor, application of the agonist DAMGO inhibited whole-cell currents through both the wild-type and the mutant channels. Prepulse facilitation was also reduced by the K1336E mutation. Likewise, the kinetic analysis of the onset and decay of facilitation showed that the mutation affects the apparent dissociation and reassociation rates of the Gβγ dimer from the channel complex. Conclusions These results suggest that the extent of G-protein-mediated inhibition is significantly reduced in the K1336E mutant CaV2.1 Ca2+ channels. This alteration would contribute to render the neuronal network hyperexcitable, possibly as a consequence of reduced presynaptic inhibition, and may help to explain some aspects of the FHM-1 pathophysiology.

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Functional correlation of ATP1A2 mutations with phenotypic spectrum: from pure hemiplegic migraine to its variant forms

The Journal of Headache and Pain ◽

10.1186/s10194-021-01309-4 ◽

2021 ◽

Vol 22 (1) ◽

Author(s):

Yingji Li ◽

Wenjing Tang ◽

Li Kang ◽

Shanshan Kong ◽

Zhao Dong ◽

...

Keyword(s):

Intellectual Disability ◽

Atpase Activity ◽

Protein Extraction ◽

Cell Voltage ◽

Familial Hemiplegic Migraine ◽

Missense Mutations ◽

Wild Type ◽

Hemiplegic Migraine ◽

Whole Cell ◽

Significant Difference

Abstract Background Mutations in ATP1A2, the gene encoding the α2 subunit of Na+/K+-ATPase, are the main cause of familial hemiplegic migraine type 2 (FHM2). The clinical presentation of FHM2 with mutations in the same gene varies from pure FHM to severe forms with epilepsy and intellectual disability, but the correlation of these symptoms with different ATP1A2 mutations is still unclear. Methods Ten ATP1A2 missense mutations were selected according to different phenotypes of FHM patients. They caused pure FHM (FHM: R65W, R202Q, R593W, G762S), FHM with epilepsy (FHME: R548C, E825K, R938P), or FHM with epilepsy and intellectual disability (FHMEI: T378N, G615R, D718N). After ouabain resistance and fluorescence modification, plasmids carrying those mutations were transiently transfected into HEK293T and HeLa cells. The biochemical functions were studied including cell survival assays, membrane protein extraction, western blotting, and Na+/K+-ATPase activity tests. The electrophysiological functions of G762S, R938P, and G615R mutations were investigated in HEK293T cells using whole-cell patch-clamp. Homology modeling was performed to determine the locational distribution of ATP1A2 mutations. Results Compared with wild-type pumps, all mutations showed a similar level of protein expression and decreased cell viability in the presence of 1 µM ouabain, and there was no significant difference among the mutant groups. The changes in Na+/K+-ATPase activity were correlated with the severity of FHM phenotypes. In the presence of 100 µM ouabain, the Na+/K+-ATPase activity was FHM > FHME > FHMEI. The ouabain-sensitive Na+/K+-ATPase activity of each mutant was significantly lower than that of the wild-type protein, and there was no significant difference among all mutant groups. Whole-cell voltage-clamp recordings in HEK293T cells showed that the ouabain-sensitive pump currents of G615R were significantly reduced, while those of G762S and R938P were comparable to those of the wild-type strain. Conclusions ATP1A2 mutations cause phenotypes ranging from pure FHM to FHM with epilepsy and intellectual disability due to varying degrees of deficits in biochemical and electrophysiological properties of Na+/K+-ATPase. Mutations associated with intellectual disability presented with severe impairment of Na+/K+-ATPase. Whether epilepsy is accompanied, or the type of epilepsy did not seem to affect the degree of impairment of pump function.

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Familial Hemiplegic Migraine Type 1 Mutations K1336E, W1684R, and V1696I Alter Cav2.1 Ca2+Channel Gating

Journal of Biological Chemistry ◽

10.1074/jbc.m408756200 ◽

2004 ◽

Vol 279 (50) ◽

pp. 51844-51850 ◽

Cited By ~ 52

Author(s):

Carmen Müllner ◽

Ludo A. M. Broos ◽

Arn M. J. M. van den Maagdenberg ◽

Jörg Striessnig

Keyword(s):

Channel Gating ◽

Brain Regions ◽

Familial Hemiplegic Migraine ◽

Mammalian Brain ◽

Β Subunit ◽

Hemiplegic Migraine ◽

Channel Function ◽

Steady State Inactivation ◽

Inward Currents

Mutations in the Cav2.1 α1-subunit of P/Q-type Ca2+channels cause human diseases, including familial hemiplegic migraine type-1 (FHM1). FHM1 mutations alter channel gating and enhanced channel activity at negative potentials appears to be a common pathogenetic mechanism. Different β-subunit isoforms (primarily β4and β3) participate in the formation of Cav2.1 channel complexes in mammalian brain. Here we investigated not only whether FHM1 mutations K1336E (KE), W1684R (WR), and V1696I (VI) can affect Cav2.1 channel function but focused on the important question whether mutation-induced changes on channel gating depend on the β-subunit isoform. Mutants were co-expressed inXenopusoocytes together with β1, β3, or β4and α2δ1subunits, and channel function was analyzed using the two-electrode voltage-clamp technique. WR shifted the voltage dependence for steady-state inactivation of Ba2+inward currents (IBa) to more negative voltages with all β-subunits tested. In contrast, a similar shift was observed for KE only when expressed with β3. All mutations promotedIBadecay during pulse trains only when expressed with β1or β3but not with β4. Enhanced decay could be explained by delayed recovery from inactivation. KE acceleratedIBainactivation only when co-expressed with β3, and VI slowed inactivation only with β1or β3. KE and WR shifted channel activation ofIBato more negative voltages. As the β-subunit composition of Cav2.1 channels varies in different brain regions, our data predict that the functional FHM1 phenotype also varies between different neurons or even within different neuronal compartments.

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