• Daniela Pietrobon
  

• Rossella Conti
  

Lab members:

• Angelita Tottene
• Dania Vecchia
• Michele Sessolo
• Andrea Urbani

Migraine is a common disabling brain disorder affecting more than 10% of the population. The primary cause of migraine lies in the brain, but the nature and mechanisms of the brain dysfunction(s) in migraine remain unclear and controversial, and drug therapy for preventing and treating migraine remains unsatisfactory for many patients. Recent findings point to cortical spreading depression (CSD) as a key player in the pathogenesis of migraine, as CSD underlies migraine aura and may also trigger the mechanisms for migraine headache. Unique insights into the pathophysiology of migraine can be gained by studying the molecular and cellular mechanisms of familial hemiplegic migraine (FHM), a monogenic subtype of migraine with aura, whose typical attacks have headache and aura symptoms similar to those of the common forms of migraine. In particular, our group studies the mechanisms of FHM type 1 (FHM1), that is caused by missense mutations in the CACNA1A gene, encoding the pore-forming subunit of a neuronal voltage-gated Ca²⁺ channel (Ca_V2.1 or P/Q-type). Ca_V2.1 channels are located in somatodendritic membranes and in presynaptic terminals throughout the brain, where they play a dominant role in initiating fast synaptic transmission. We have shown that:
1) FHM1 mutations produce gain-of-function of human recombinant Ca_V2.1 channels, mainly due to a shift of channel activation to more negative voltages and an increase of the open probability and single channel influx over a broad voltage range;
2) knockin (KI) mice carrying FHM1 mutations (either the mild R192Q or the severe S218L) show an increased P/Q-type Ca²⁺ current in cerebellar and cortical pyramidal neurons, and an increased strength of cortical excitatory synaptic transmission due to increased action potential-evoked Ca²⁺ influx and increased probability of glutamate release at pyramidal cell synapses; (Fig 1)
3) both the induction and the propagation of experimental CSD, elicited either by electrical stimulation in vivo or high KCl in acute slices of sensory cortex, are facilitated in FHM1 KI mice. The exten t of CSD facilitation correlates with the severity of the clinical phenotype of the FHM1 mutations.

To gain insights into the unknown mechanisms that lead to CSD susceptibility and initiate migraine attacks in patients, our current work is concentrated on the investigation of the cortical mechanisms that produce facilitation of CSD in FHM1 mouse models. Patch-clamp recordings on cortical neurons in microculture and in acute brain slices and paired patch-clamp recordings under IR-DIC microscopy on connected neurons in acute thalamocortical slices are used to study cortical excitatory and inhibitory neurotransmission and cortical network excitability in R192Q and S218L KI mice. Using these methods, we have recently shown that in contrast with the enhanced excitatory synaptic transmission at cortical pyramidal cell synapses, inhibitory synaptic transmission at fast-spiking interneuron synapses is not altered in R192Q KI mice (Fig 2).

The in-vitro model of CSD, recently developed by our group, is used to study the mechanisms of initiation and propagation of CSD and to investigate the role of neuron-astrocyte signaling in these mechanisms. Using this system we have recently obtained direct evidence of a causative link between enhanced glutamate release at pyramidal cell synapses and facilitation of experimental CSD in R192Q KI mice (Fig 3). In the near future the electrophysiological measurements in cortical slices will be complemented by Ca²⁺ imaging experiments to follow simultaneously the activity of many cells and to investigate directly whether FHM KI mice show alterations in the spatiotemporal patterns of network activity. An important part of this project is to confirm and expand in the living brain the findings obtained in acute cortical slices.

Fig. 1

Fig. 2

Fig. 3