No abstract

Slit- and Trk-like (Slitrks) are a six-member family of synapse organizers that control excitatory and inhibitory synapse formation by forming trans-synaptic adhesions with LAR receptor protein tyrosine phosphatases (PTPs). Intriguingly, genetic mutations of Slitrks have been associated with a multitude of neuropsychiatric disorders. However, nothing is known about the neuronal and synaptic consequences of these mutations. Here, we report the structural and functional effects on synapses of various rare de novo mutations identified in patients with schizophrenia or Tourette syndrome. A number of single amino acid substitutions in Slitrk1 (N400I or T418S) or Slitrk4 (V206I or I578V) reduced their surface expression levels. These substitutions impaired glycosylation of Slitrks expressed in HEK293T cells, caused retention of Slitrks in the endoplasmic reticulum and cis-Golgi compartment in COS-7 cells and neurons, and abolished Slitrk binding to PTPδ. Furthermore, these substitutions eliminated the synapse-inducing activity of Slitrks, abolishing their functional effects on synapse density in cultured neurons. Strikingly, a valine-to-methionine mutation in Slitrk2 (V89M) compromised synapse formation activity in cultured neuron, without affecting surface transport, expression, or synapse-inducing activity in coculture assays. Similar deleterious effects were observed upon introduction of the corresponding valine-to-methionine mutation into Slitrk1 (V85M), suggesting that this conserved valine residue plays a key role in maintaining the synaptic functions of Slitrks. Collectively, these data indicate that inactivation of distinct cellular mechanisms caused by specific Slitrk dysfunctions may underlie Slitrk-associated neuropsychiatric disorders in humans, and provide a robust cellular readout for the development of knowledge-based therapies.

Abstract-Mechanosensitive cation channels may be involved in the development of the myogenic tone of arteries. The molecular identity of these channels is not clear, but transient receptor potential channels (TRPCs) are good candidates.In the present study, we searched for mechanosensitive channels at the single-channel level in arterial smooth muscle cells using the patch-clamp technique and investigated the channel properties in the light of properties of TRPCs. With 140 mmol/L CsCl in the pipette solution, application of negative pressures to the back of the pipette induced the activation of channels the open probability of which increased with the amount of negative pressure. The current-voltage relationship was linear in symmetrical ionic conditions, and the single-channel conductances for Cs ϩ , K ϩ , and Na ϩ were 30, 36, and 27 pS, respectively. When NMDG ϩ was substituted for Cs ϩ in the pipette solution, inward currents were abolished, whereas outward currents remained active, indicating that the channels were nonselective to cations. The channel activity was blocked by intracellular Gd 3ϩ and 4,4Ј-diisothiocyanatostilbene-2,2Ј-disulfonic acid and increased by diacylglycerol and by cyclopiazonic acid. Phospholipase C inhibitor (U73122) inhibited not only channel activity but also the development of myogenic tone induced by stretching of the basilar arteries. These results suggest that the ion channel responsible for the development of myogenic tone is the 30-pS mechanosensitive cation channel that exhibits properties similar to those of TRPCs. Ⅲ myogenic tone Ⅲ phospholipase C T he arterial wall is continuously exposed to mechanical stimulation such as shear stress and luminal pressure. It is well known that such mechanical stimulation causes various biological responses in the vascular wall. [1][2][3][4] One of the important responses is the development of myogenic tone: a state of partial contraction that is dependent on the level of intraluminal pressure. This phenomenon was first described by Bayliss 5 100 years ago, but the exact signal transduction pathways are still unclear.Pressure-induced depolarization and the increase in intracellular Ca 2ϩ concentration are associated with the development of myogenic tone when the transmural pressure is elevated to 40 to 60 mm Hg. 6 Therefore, mechanosensitive ion channels, especially nonselective cation (NSC) channels, are thought to transduce the change in transmural pressure into membrane depolarization, and the activation of wholecell cation currents in response to longitudinal stretch of isolated smooth muscle cells has been demonstrated by several authors. 2,3,7 However, the identity of the mechanosensitive NSC channels in smooth muscle cells remains unclear.Transient receptor potential channels (TRPCs) are good candidates for this role in arterial smooth muscle, because they are expressed in vascular smooth muscle cells and they exhibit many of the biological properties of vascular cation currents. 8 -11 This hypothesis was tested recently by Welsh ...