Whole cell patch-clamp recordings of dissociated ommatidia from Sh14 flies show a similar leftward shift of the I/V curve in response to green light stimulation as do recordings of wild-type dissociated ommatidia.
Heterozygous flies show a rapid leg-shaking phenotype when under ether anesthesia.
At low Ca2+ levels (0.1mM), the excitatory junctional potential (ejp) elicited in the muscle at the neuromuscular junction has increased amplitude and duration in mutant larvae compared to wild type.
Sh14 flies do not show a significantly shortened lifespan. At day 35, the brains of these flies show intermittent pathology.
Electrical current recordings from Sh14 mutant photoreceptors show the transient K+ current is eliminated, but the slowly inactivating current is intact in these mutants.
Mutant flies exhibit a decrease in the daily sleep amount. This is mainly due to a decrease in the duration of sleep episodes rather than in their number. This phenotype is very sensitive to background modifiers.
Mutant muscles (ventral lateral longitudinal fiber 6 of segments A2 and A3 have been tested) lack the prominent A-type inactivating outward K+ current. Dissociated mushroom body intrinsic neurons do not show a dramatic modification of the whole-cell outward K+ current profile or a significant decrease in the current density compared to wild type. Two neuronal populations are present in preparations of wild-type dissociated mushroom body intrinsic neurons; a major population (approximately 72%) having half-inactivation voltages of approximately -78mV, and a minor one having more depolarised half-inactivation voltage values. This second population is absent in preparations of Sh14 dissociated mushroom body intrinsic neurons.
Mutant adults exhibit both leg shaking and wing scissoring under ether anaesthetization. This behaviour persists in severed legs.
Sh14 photoreceptors are grossly morphology normal. Sh14 photoreceptors are unable to follow sequences of rapid transients, unlike wild-type receptors. For example, light intensity fluctuations of increasing intensity are encoded by increasingly frequent responses in the wild-type photoreceptor, while Sh14 responses approach a plateau of depolarisations at lower stimulus levels, compressing their responses to bright inputs. Mutant photoreceptors produce large, fast voltage responses to large light intensity increases after dim periods, which are reduced in wild-type photoreceptors. During sustained periods of bright light Sh14 receptors produce smaller responses than wild-type. Additionally, Sh14 voltage responses repolarise more rapidly than wild-type voltage responses.
Voltage responses of Sh14 photoreceptors to naturalistic stimuli are altered compared to wild-type. In particular, many of the transient responses to naturalistic stimuli in Sh14 are distorted, and the spread of the signal over the available voltage range is reduced in Sh14 photoreceptors. Sh14 photoreceptors have large-amplitude, shorter time-to-peak, and shorter half-width linear impulse responses than wild-type.
When presented with sequences of dynamically modulated light with a mean contrast of 0.32, close to that of natural sceneries, over the range of intensities to which photoreceptors are normally exposed, the light induced current, quantum efficiency and macroscopic kinetics are unaffected in Sh14 photoreceptors. The photoreceptor signal response in reduced in mutant animals, at all background intensities. Mutant flies also have reduced information capacity of photoreceptors at all but the dimmest light levels. The information loss is greatest over the lower frequency range of the spectrum (1-50Hz). Under light adapted conditions the N(f) of mutant photoreceptors are similar to wild-type. Mutant photoreceptor show a reduced contrast gain and broadened bandwidth with increasing mean light intensity. Sh14 responses contain more high-frequency signals. Mutant photoreceptors have reduced resistances compared to wild-type. Current pulse experiments show that the steady-state conductance of mutant photoreceptors behave as would be expected of a wild-type membrane without Sh, with no further voltage-dependent conductances.
Sh14 flies show normal osmotic stress sensitivity on food medium containing NaCl.
The seizure threshold following short wavetrains of high-frequency electrical stimuli is increased in mutant flies compared to controls; seizures cannot be evoked with the standard 300ms high-frequency stimuli at 100V, but can be evoked using 400ms high-frequency stimuli. Under these conditions, the seizure threshold of the mutant flies is 83.8 +/- 12.8 V.
The threshold for activation of the giant fiber in mutant animals following single stimulus pulses (0.2ms duration, 0.5Hz) is not significantly different from that of wild type.
The giant fiber following frequency (the maximum stimulation frequency that the giant fiber pathway can reliably follow) is greatly reduced compared to wild type in mutant animals.
Homozygous Sh14 females exhibit a marked increase in sensitivity to halothane in an electrophysiologically monitored escape response. Specifically, halothane EC[] is increased in Sh14 stocks by 85% over that in wild-type control females. Heterozygous Sh14 females exhibit a halothane potency that is intermediate between the homozygous mutant and controls. Male Sh9 mutants produce a modest increase in EC[] (35% compared to Canton-S flies).
The resting potentials of mutant dorsolongitudinal muscles does not differ significantly from wild type at 22 or 12oC.
Mutant flies show an increased sensitivity to halothane in an inebriometer assay compared to control flies, having a higher Response Index of 0.85 +/- 0.09 after 12 minutes of exposure to halothane.
IA and IK currents are present in cell cultures of midline neurons prepared from Sh14 mutants.
Sh14 flies treated with sodium valproate (NaVP) show a reduction in body weight, as is seen in wild-type flies. Treatment of Sh14 flies with NaVP causes an increase in mortality as is seen in wild-type flies.
slo4 partially suppresses the synaptic transmission defects (eliminates the repetitive firing of motor axons).
Homozygotes show increased sensitivity to halothane, chloroform and trichloroethylene in an inebriometer assay (an assay of geotactic and postural behaviour) compared to wild-type flies. Decapitated flies lose the halothane sensitive phenotype.
More sensitive than the wild-type to the knock-down effect of acute γ-irradiation.
The delivery of an electrical buzz (50-400 msec) to the brain has no significant effect on Sh14 mutant flies.
Two-electrode voltage clamp technique is used to measure end-plate currents in larval neuromuscular junctions. Currents are four fold larger than wild type, lack post-tetanic potentiation (PTP) but could undergo facilitation. PTPs could be restored by addition of Cd2+.
Vigorous and continuous vibration of all appendages. Double mutant combinations with tta1 show slight suppression of the Sh phenotype. Sh phenotype is enhanced by the presence of one extra copy of tta.
Completely removes the fast voltage-gated potassium current in muscle during all stages of development (FBrf0043054). Muscles show abnormal transmission characterised by increased transmitter release and prolonged excitatory junction potentials (EJPs) (FBrf0043046). eag1 Sh14 double mutant embryos show greatly increased synaptic activity, both the burst frequency and the mean excitatory junction current (EJC) amplitude are increased during synaptogenesis. The synaptic morphology of the double mutant embryonic NMJs raised at the restrictive temperature are not significantly different from wild type, at the light microscope level. These hyperactive synapses expand to occupy a larger area of the muscle surface relative to wild type causing decreased synaptic density (the branch and bouton numbers are not altered).
eag1, Sh14 double mutants have normal grooming behavior.
Flies show leg-shaking under anaesthesia. The action potential of the dorsal longitudinal muscle often appears abnormal in Sh14 mutants and Sh14 eag1 double mutants. Increases the refractory period and lowers the following frequency of the giant fibre response of the dorsal longitudinal muscle pathway and of the tergotrochanteral muscle pathway. eag1 Sh14 double mutant flies show spontaneous activity of the dorsal longitudinal and dorsoventral indirect flight muscles when at rest. 86% of eag1 Sh14 flies have a wings-down phenotype.
Sh14 and DAP (3,4-diaminopyridine) cause the same increase in synaptic delay and the same reduction in the slope of onset phase of electrotonically evoked synaptic current in slo mutant third instar larvae.
Leg shaking while under ether anaesthesia.
The mutants effects can be potentiated by mutations in dnc and the enhancement can be counterbalanced by rut1. The effect of extreme hyperexcitability in eag Sh double mutants cannot be potentiated by mutations in dnc.
Heterozygotes show ether induced leg shaking. Hemizygous males show a reduced preference for sucrose compared to wild-type in feeding preference tests, probably due to a shift in the threshold of detection. Flies show no attraction to 100mM NaCl and an increased tolerance to 0.5M NaCl and 0.2M KCl compared to wild-type. The increased tolerance to 0.5M NaCl and 0.2M KCl is due to an increase in the threshold of repulsion. The firing patterns of the labellar chemosensory neurons in response to sucrose, NaCl and KCl are normal.
Muscle fibres and photoreceptors have no detectable IA (rapidly inactivating A current).
Presynaptic action potential decay time is considerably prolonged in Sh14 third instar neuromuscular junctions compared to wild-type.
The leg shaking phenotype of Sh14 is enhanced by Dp(1;4)r+l. eag1 Sh14 double mutant larvae show an increased number of tertiary and quarternary axonal branches over muscles 12 and 13. The number, and in some cases density, of varicosities in the neurites is also increased.
Flies manifest chronic vibration of their appendages as well as abnormal action potentials.
IA is completely eliminated in homozygous third larval instar muscles. IA is much lower than predicted by simple gene-dosage dependence in Sh14/+, Sh14/Sh21, Sh14/Sh9 and Sh14/Sh5 flies.
The voltage-activated fast K+ current (IA) is eliminated in larval muscle fibres.
Heterozygous flies show rapid-leg shaking under ether anesthesia.
A-type current is completely absent.
Excitatory junction potentials are prolonged even at very low external Ca2+ concentrations. eag1 interacts synergistically with Sh14 in double mutants to produce extremely prolonged neuromuscular transmission and spontaneous firing of the motor axons.
Ether-dependent leg shaking and wing scissoring. Chloroform, ethyl acetate and carbon dioxide etherization does not elicit shaking behavior, though nitrogen and triethylkamine etherization does. Unetherized, older flies show uncoordinated walking behavior, and stand quivering on the bottom of the culture bottle. In the larval neuromuscular junction preparation, repetetive firing of the motor axons is associated with a prolonged release of neurotransmitter at the nmj producing a prolonged, large ejp.
abnormal leg shaking under ether anesthesia; abnormal A-type potassium currents in larval muscle and/or pupal flight muscle; abnormal action potentials in the adult cervical giant fiber; abnormal synaptic transmission at the larval neuromuscular junction and multiple firing of larval motoneurons.