Altered Cerebellar Short-Term Plasticity but No Change in Postsynaptic AMPA-Type Glutamate Receptors in a Mouse Model of Juvenile Batten Disease

Juvenile Batten disease is the most common progressive neurodegenerative disorder of childhood. It is associated with mutations in the CLN3 gene, causing loss of function of CLN3 protein and degeneration of cerebellar and retinal neurons. It has been proposed that changes in granule cell AMPA-type glutamate receptors (AMPARs) contribute to the cerebellar dysfunction. In this study we compared AMPAR properties and synaptic transmission in cerebellar granule cells from wild-type and Cln3 knockout mice. In Cln3Δex1–6 cells the amplitude of AMPA-evoked whole-cell currents was unchanged. Similarly, we found no change in the amplitude, kinetics, or rectification of synaptic currents evoked by individual quanta, or in their underlying single-channel conductance. We found no change in cerebellar expression of GluA2 or GluA4 protein. By contrast, we observed a reduced number of quantal events following mossy-fiber stimulation in Sr2+, altered short-term plasticity in conditions of reduced extracellular Ca2+, and reduced mossy fiber vesicle number. Thus, while our results suggest early presynaptic changes in the Cln3Δex1–6 mouse model of juvenile Batten disease, they reveal no evidence for altered postsynaptic AMPARs

Enhanced functional detection of synaptic calcium-permeable AMPA receptors using intracellular NASPM

Calcium-permeable AMPA-type glutamate receptors (CP-AMPARs) contribute to many forms of synaptic plasticity and pathology. They can be distinguished from GluA2-containing calcium-impermeable AMPARs by the inward rectification of their currents, which reflects voltage-dependent channel block by intracellular spermine. However, the efficacy of this weakly permeant blocker is differentially altered by the presence of AMPAR auxiliary subunits - including transmembrane AMPAR regulatory proteins, cornichons and GSG1L - which are widely expressed in neurons and glia. This complicates the interpretation of rectification as a measure of CP-AMPAR expression. Here we show that inclusion of the spider toxin analogue 1‑naphthylacetyl spermine (NASPM) in the intracellular solution results in complete block of GluA1-mediated outward currents irrespective of the type of associated auxiliary subunit. In neurons from GluA2-knockout mice expressing only CP-AMPARs, intracellular NASPM, unlike spermine, completely blocks outward synaptic currents. Thus, our results identify a functional measure of CP-AMPARs, that is unaffected by their auxiliary subunit content

TARP γ-7 selectively enhances synaptic expression of calcium-permeable AMPARs

Regulation of calcium-permeable AMPA receptors (CP-AMPARs) is crucial in normal synaptic function and neurological disease states. Although transmembrane AMPAR regulatory proteins (TARPs) such as stargazin (γ-2) modulate the properties of calcium-impermeable AMPARs (CI-AMPARs) and promote their synaptic targeting, the TARP-specific rules governing CP-AMPAR synaptic trafficking remain unclear. We used RNA interference to manipulate AMPAR-subunit and TARP expression in γ-2–lacking stargazer cerebellar granule cells—the classic model of TARP deficiency. We found that TARP γ-7 selectively enhanced the synaptic expression of CP-AMPARs and suppressed CI-AMPARs, identifying a pivotal role of γ-7 in regulating the prevalence of CP-AMPARs. In the absence of associated TARPs, both CP-AMPARs and CI-AMPARs were able to localize to synapses and mediate transmission, although their properties were altered. Our results also establish that TARPed synaptic receptors in granule cells require both γ-2 and γ-7 and reveal an unexpected basis for the loss of AMPAR-mediated transmission in stargazer mice