Alk^ts/Alk¹ flies raised at 18[o] do not have significant changes in body length.

When sleep is measured using single beam monitors at the permissive temperature (18[o]C), Alk^ts/Alk¹ females have a mild but significant increase in total sleep compared to controls (no significant changes in males); acute inhibition of Alk at the restrictive temperature (29[o]C) in Alk^ts/Alk¹ flies leads to significant increases in sleep amount compared to controls (no effect in Alk^ts/+ females, a small but significant increase in males).

When sleep is measured using multi-beam monitors at 18[o]C, Alk^ts/Alk¹ flies have similar total sleep amount as controls, but at 29[o]C show significant total sleep increases (both day and night). At 29[o]C both Alk^ts/Alk¹ and Alk¹/+ females have significant decreases in waking activity and show normal geotaxis activity.

Alk^ts/Alk¹ raised at 18[o]C maintain locomotor rhythms at 29[o]C in darkness.

Alk^ts fails to complement Alk¹ lethality at 29[o]C and shows a mutant midgut phenotype. Alk¹/Alk^ts animals show larval lethality at 25[o]C.

Alk¹/Alk^ts animals can be effectively maintained to third instar or pupal stages by initial rearing at 18[o]C for 48-72 hours from egg laying, followed by subsequent rearing at 25[o]C to the third larval instar. Excitatory junctional current (EJC) amplitude is increased by 210% compared to controls in Alk¹/Alk^ts third instar larvae raised under these conditions.

Homozygous Alk¹ mutant longitudinal visceral muscle (LVM) founder cells in stage 12 embryos migrate normally towards the trunk visceral mesoderm (TVM) and very few dying cells are seen. At stage 13 the front migrating cells reach the anterior end of the trunk visceral mesoderm as in wild type. However during late stage 13 the migration becomes disordered and progressive founder cell death is seen.

Alk¹ heterozygous pupae are normal in size.

The central nervous system of Alk¹ mutants appears morphologically normal in terms of segmental nerve branching and segmental muscle patterning. Neuropil synaptic differentiation appears normal in these mutants, with comparable labeling intensity, density and distribution of presynaptic and postsynaptic labels.

Alk¹ mutant neuromuscular junctions appear correctly and stereotypically formed and elaborated, with no examples of muscle innervation failure or synaptic targeting errors. mutant presynaptic active zones appear normally formed and with wild-type size.

Homozygous Alk¹ mutants exhibit reduced hatching and mutant larval movement is typically sluggish and highly limited. Locomotory movement in Alk¹ mutants is decreased to 35% of the control level, characterised by slower contractions and extended pauses. Severely impaired and motionless larvae remain capable of briefly resuming movement when stimulated, suggesting a defect in the central circuit output driving locomotion.

Alk¹ mutant neuromuscular junctions exhibit visible glutamate-driven muscle contractions, and consistently large and robust postsynaptic current amplitudes compared to controls.

In Alk¹ mutant neuromuscular junctions, no large or patterned EJCs are recorded (maximum amplitude 429nA), and overall EJC frequency is below 1Hz in 80% of active cells.

R cells, target neurons and glia are normal in Alk¹ eye mosaic clones.

Third instar larvae that have Alk function removed in the lamina target neurons, by the presence of Alk¹ clones, show normal R cell guidance. Additionally, adults with eyes made mutant for Alk¹ by the presence of clones show no R cell guidance defects.

Adult mosaic Alk¹ animals, in which lamina target neurons are mutant for Alk¹, show R8 axon guidance defects. R8 cells exhibit thickened terminals and frequently fail to terminate in their appropriate layer. These axons either project into the R7 layer (7%), stop at the distal border of the medulla neuropil (12%), or project to neighbouring columns (9%). The vast majority of R7 terminals target normally but a small proportion extend into an adjacent medulla column.

At the pupal stage, animals with lamina target neuron Alk¹ clones exhibit normal R cell projections up to 40 hours after puparium formation. At around 55 hours, R cell projection defects become apparent. These include thickened terminals, fasciculation with neighbouring columns and stalling of the axons at the distal medulla neuropil border ot projection into an adjacent column.

In adults carrying Alk¹ clones in the lamina target neurons, the array of cartridges consisting of R1-R6 terminals in the lamina is disrupted and the size and shape of lamina cartridges is variable. Cartridges containing homozygous mutant target lamina are likely to be distributed randomly throughout the lamina.

The visceral mesoderm is disrupted in stage 14 Alk¹ embryos.

Alk¹ homozygous larvae do not ingest food, and lack discernible intestinal structures. In stage 13 Alk¹ homozygous embryos, the visceral mesoderm is disorganized. At stage 11, the earliest stage at which a mutant phenotype in the visceral mesoderm can clearly be observed, these embryos lack muscle founder cells. However, myoblasts do form in the visceral mesoderm, and go on to contribute to somatic muscle (i.e.- they are fusion competent.)

55% of Alk¹/+ mutants die as embryos and the rest die as first-instar larvae.

Alk¹ first-instar larvae are unable to eat as the visceral mesoderm is severely disrupted and no functional midgut is formed.

Alk¹/Alk[+] is a suppressor | partially of abnormal size | pupal stage phenotype of Nf1^E2

Alk¹/Alk[+] is a suppressor | partially of abnormal learning | adult stage phenotype of Nf1^E2