title: Loss of β2-laminin alters calcium sensitivity and voltage-gated calcium channel maturation of neurotransmission at the neuromuscular junction.
abstract: Neuromuscular junctions from β2-laminin-deficient mice exhibit lower levels of calcium sensitivity. Loss of β2-laminin leads to a failure in switching from N- to P/Q-type voltage-gated calcium channel (VGCC)-mediated transmitter release that normally occurs with neuromuscular junction maturation. The motor nerve terminals from β2-laminin-deficient mice fail to up-regulate the expression of P/Q-type VGCCs clusters and down-regulate N-type VGCCs clusters, as they mature. There is decreased co-localisation of presynaptic specialisations in β2-laminin-deficient neuromuscular junctions as a consequence of lesser P/Q-type VGCC expression. These findings support the idea that β2-laminin is critical in the organisation and maintenance of active zones at the neuromuscular junction via its interaction with P/Q-type VGCCs, which aid in stabilisation of the synapse. β2-laminin is a key mediator in the differentiation and formation of the skeletal neuromuscular junction. Loss of β2-laminin results in significant structural and functional aberrations such as decreased number of active zones and reduced spontaneous release of transmitter. In vitro β2-laminin has been shown to bind directly to the pore forming subunit of P/Q-type voltage-gated calcium channels (VGCCs). Neurotransmission is initially mediated by N-type VGCCs, but by postnatal day 18 switches to P/Q-type VGCC dominance. The present study investigated the changes in neurotransmission during the switch from N- to P/Q-type VGCC-mediated transmitter release at β2-laminin-deficient junctions. Analysis of the relationship between quantal content and extracellular calcium concentrations demonstrated a decrease in the calcium sensitivity, but no change in calcium dependence at β2-laminin-deficient junctions. Electrophysiological studies on VGCC sub-types involved in transmitter release indicate N-type VGCCs remain the primary mediator of transmitter release at matured β2-laminin-deficient junctions. Immunohistochemical analyses displayed irregularly shaped and immature β2-laminin-deficient neuromuscular junctions when compared to matured wild-type junctions. β2-laminin-deficient junctions also maintained the presence of N-type VGCC clustering within the presynaptic membrane, which supported the functional findings of the present study. We conclude that β2-laminin is a key regulator in development of the NMJ, with its loss resulting in reduced transmitter release due to decreased calcium sensitivity stemming from a failure to switch from N- to P/Q-type VGCC-mediated synaptic transmission.
authors.affiliation: School of Biomedical Sciences, The University of Queensland, St Lucia, 4067, Australia.
authors.forename: Kirat K
authors.initials: KK
authors.lastname: Chand
authors.forename: Kah Meng
authors.initials: KM
authors.lastname: Lee
authors.forename: Mitja P
authors.initials: MP
authors.lastname: Schenning
authors.forename: Nickolas A
authors.initials: NA
authors.lastname: Lavidis
authors.forename: Peter G
authors.initials: PG
authors.lastname: Noakes
mesh_terms: D000818:Animals; D002118:Calcium / Q000502:physiology; D015220:Calcium Channels / Q000502:physiology*; D005260:Female; D007797:Laminin / Q000235:genetics / Q000502:physiology*; D008297:Male; D018345:Mice, Knockout; D009469:Neuromuscular Junction / Q000502:physiology*; D010791:Phrenic Nerve / Q000502:physiology*; D009435:Synaptic Transmission / Q000502:physiology*
chemical_list: D015220:Calcium Channels; D007797:Laminin; C058783:laminin beta2; D002118:Calcium