1. Spike frequency adaptation mediates looming stimulus selectivity in a collision-detecting neuron Peron and Gabbiani – Nature Neuroscience, March 2009 In Houston at Rice and Baylor

2. Terminology Spike Frequency Adaptation (SFA) - firing frequency to decrease during a train of action potentials. Translating Stimuli Looming Stimuli X Y = Y dependent X Channels K ca = Calcium dependent Potassium Channels

3. Escape Behavior and Locomotion Escape Movement occurs at angle threshold (when triangle is sufficiently large enough to cover the needed number of retinal receptors) Retinal neurons - LGMD interneurons – Lobula Giant Movement Detector To DCMD – Descending Contralateral Movement Detector To muscles. Usually a 1:1 correspondence from LGMD to DCMD.

4. Methods - Differences In vivo - Locus chosen because had to present visual stimuli in vivo. Larger input currents, 10+ nA (e-9) compared to pico (e-12) amps (our lab). Use Matlab instead of Igor. Locus saline instead of ACSF. Locus – a model organism

5. Methods - Alan Lloyd Hodgkin Goldman-Hodgkin-Katz Hodgkin–Huxley model - 1963 Nobel Prize with Huxley - model that describes how action potentials in neurons are initiated and propagated Cm = capaticance – lipid bilayer Gn and Gl = liner and non-linear ion flow Ip = current source = Na/K Pump

6. The Locus escape mechanism is activated in response to looming stimuli but not translating stimuli. How does this occur?

7. LGMD responds to looming stimuli Maximum frequency in dynamic range - 27spikes/sec versus 300spikes/sec. Current was 10nA for locus.

8. LGMD Response - Looming versus Translating 300/sec versus 50spikes/sec (6x) with lower sustained firing rates When looking at last.4 sec of the 1 sec pulse, a difference of 100 spikes looming versus 10 spikes for translating (10x) Translating stimuli show spike frequency adaptation while looming stimuli do not. SFA leading to repressed translational stimuli response Looming correlated with motor response. Correlation is not causation.

9. SK-Like Potassium conductance mediates the spike-frequency adaptation in translating stimuli SK - Small conductance K+ channel – activated by calcium – too slow to contribute to repolarization but SK currents contributes to medium after-hyperpolarization. How was this conclusion reached? - BAPTA

10. BAPTA addition Calcium Chelator BAPTA – Forms a chelate complex with calcium - sequesters calcium Calcium Channel blocker Cadmium (Cd2+) Chelator sequesters calcium while Cadmium prevents Calcium entry RESULT : Post BAPTA administration - steady translating stimuli frequency, No adaptation SFA is calcium-dependent

11. Singling out SK from the K ca Class K ca – SK (small-conductance), BK, IK (=I sAHP? ) Use of Charybdotoxin – blocks BK and IK but not SK – Reduced spike frequency and broadened the action potential but no effect on final spike frequency, SFA still observed. However, Apamin – traditional SK antagonist - and other SK blockers had no effect on SFA. SFA was still present.

12. SK-like Conductance responsible for SFA SFA seems to be mediated by SK – confirmed calcium dependence and all other K ca ruled out. But - aunique SK showing “nontraditional properties.” “SK-like” – something new?

13. A side investigation into BAPTA BAPTA abolishes the 1:1 LGMD – DCMD ratio. Chemical Synapse Review: Calcium diffuses into synaptic terminal in response to action potential repolarization leading to release of neurotransmitters from calcium sensitive vesicles. Shows BAPTA is able to diffuse throughout the LGMD – all the way to the LGMD –DCMD electrical-chemical synapse. Hypothesis Reconfirmation - Increased firing frequency observed in response to translating stimuli post BAPTA treatment.

14. BAPTA effect on Looming Stimuli “Overall course of looming stimuli response was nearly unaffected” Angle threshold for number of receptor activated by looming stimuli was not changed.

15. Spike Initiation Zone = Axon Hillock? Proximal conductance has more influence than distal conductance. SFA maximum inhibitory effect when located as close as possible to Spike Initiation Zone where input currents are summated at axon hillock. SK localized to points close to SIZ through Calcium Indicator Oregon Green BAPTA-1. Because BAPTA complexes with Calcium, can see where Calcium is localized. Fluorescence observed near SIZ.

16. BAPTA and the time constant Addition of BAPTA resulted in a decreased time constant for calcium In other words – there is a faster decline in free calcium when BAPTA is around. Time constant for Calcium needed to be obtained for use in mathematical calculations in MATLAB to confirm results

17. Mathematical Confirmation based on the Hodgkin–Huxley model Based Gn and Gl on SK-Like K ca and modified for the morphology of the LGMD and Calcium localization. Also add BAPTA constraints – Plug into MATLAB Result: SK-Like K ca conductance can almost perfectly account for the observed translational adaptation.

18. Important Results End result – New mechanism for neuron stimuli tuning – SK Channel mediated SFA in the LGMD is responsible for repression of translational stimuli. Because SK channels are responsible for medium AHP, Increasing numbers of activated SK channels gives a longer after hyperpolarization – resulting in a longer period of hyperpolarization – end result is a lowered firing frequency through adaptation. Locus achieves looming stimuli sensitivity through translational stimuli repression.

19. Remaining Questions – Why were the “SK- Like” channels unaffected by apamin and or other known SK blockers? SK channel mutation or new channel class?