A cGMP-dependent protein kinase gene, foraging, modifies habituation-like response decrement of the giant fiber escape circuit in Drosophila.

The Drosophila giant fiber jump-and-flight escape response is a model for genetic analysis of both the physiology and the plasticity of a sensorimotor behavioral pathway. We previously established the electrically induced giant fiber response in intact tethered flies as a model for habituation, a form of nonassociative learning. Here, we show that the rate of stimulus-dependent response decrement of this neural pathway in a habituation protocol is correlated with PKG (cGMP-Dependent Protein Kinase) activity and foraging behavior. We assayed response decrement for natural and mutant rover and sitter alleles of the foraging (for) gene that encodes a Drosophila PKG. Rover larvae and adults, which have higher PKG activities, travel significantly farther while foraging than sitters with lower PKG activities. Response decrement was most rapid in genotypes previously shown to have low PKG activities and sitter-like foraging behavior. We also found differences in spontaneous recovery (the reversal of response decrement during a rest from stimulation) and a dishabituation-like phenomenon (the reversal of response decrement evoked by a novel stimulus). This electrophysiological study in an intact animal preparation provides one of the first direct demonstrations that PKG can affect plasticity in a simple learning paradigm. It increases our understanding of the complex interplay of factors that can modulate the sensitivity of the giant fiber escape response, and it defines a new adult-stage phenotype of the foraging locus. Finally, these results show that behaviorally relevant neural plasticity in an identified circuit can be influenced by a single-locus genetic polymorphism existing in a natural population of Drosophila.

Experience-dependent modification of ultrasound auditory processing in a cricket escape response.

The ultrasound acoustic startle response (ASR) of crickets (Teleogryllus oceanicus) is a defense against echolocating bats. The ASR to a test pulse can be habituated by a train of ultrasound prepulses. We found that this conditioning paradigm modified both the gain and the lateral direction of the startle response. Habituation reduced the slope of the intensity/response relationship but did not alter stimulus threshold, so habituation extended the dynamic range of the ASR to higher stimulus intensities. Prepulses from the side (90 degrees or 270 degrees azimuth) had a priming effect upon the lateral direction of the ASR, increasing the likelihood that test pulses from the front (between -22 degrees and +22 degrees ) would evoke responses towards the same side as prepulse-induced responses. The plasticity revealed by these experiments could alter the efficacy of the ASR as an escape response and might indicate experience-dependent modification of auditory perception. We also examined stimulus control of habituation by prepulse intensity or direction. Only suprathreshold prepulses induced habituation. Prepulses from one side habituated the responses to test pulses from either the ipsilateral or contralateral side, but habituation was strongest for the prepulse-ipsilateral side. We suggest that habituation of the ASR occurs in the brain, after the point in the pathway where the threshold is mediated, and that directional priming results from a second process of plasticity distinct from that underlying habituation. These inferences bring us a step closer to identifying the neural substrates of plasticity in the ASR pathway.

Genetic dissection of functional contributions of specific potassium channel subunits in habituation of an escape circuit in Drosophila.

Potassium channels have been implicated in central roles in activity-dependent neural plasticity. The giant fiber escape pathway of Drosophila has been established as a model for analyzing habituation and its modification by memory mutations in an identified circuit. Several genes in Drosophila encoding K+ channel subunits have been characterized, permitting examination of the contributions of specific channel subunits to simple conditioning in an identified circuit that is amenable to genetic analysis. Our results show that mutations altering each of four K+ channel subunits (Sh, slo, eag, and Hk) have distinct effects on habituation at least as strong as those of dunce and rutabaga, memory mutants with defective cAMP metabolism (). Habituation, spontaneous recovery, and dishabituation of the electrically stimulated long-latency giant fiber pathway response were shown in each mutant type. Mutations of Sh (voltage-gated) and slo (Ca2+-gated) subunits enhanced and slowed habituation, respectively. However, mutations of eag and Hk subunits, which confer K+-current modulation, had even more extreme phenotypes, again enhancing and slowing habituation, respectively. In double mutants, Sh mutations moderated the strong phenotypes of eag and Hk, suggesting that their modulatory functions are best expressed in the presence of intact Sh subunits. Nonactivity-dependent responses (refractory period and latency) at two stages of the circuit were altered only in some mutants and do not account for modifications of habituation. Furthermore, failures of the long-latency response during habituation, which normally occur in labile connections in the brain, could be induced in the thoracic circuit stage in Hk mutants. Our work indicates that different K+ channel subunits play distinct roles in activity-dependent neural plasticity and thus can be incorporated along with second messenger "memory" loci to enrich the genetic analysis of learning and memory.

Altered habituation of an identified escape circuit in Drosophila memory mutants.

Genetic approaches in Drosophila have advanced our understanding of the molecular mechanisms of different forms of learning, including habituation, but relevant neural components have not been explored. We show that a well defined neural circuit that underlies an escape response can be habituated, providing for the first time excellent opportunities for studying physiological parameters of learning in a functional circuit in the fly. Compared with other forms of conditioning, relatively little is known of the physiological mechanisms of habituation. The giant fiber pathway mediates a jump-and-flight escape response to visual stimuli. The jump may also be triggered electrically at multiple sites in the tethered fly. This response shows parameters of habituation, including frequency-dependent decline in responsiveness, spontaneous recovery, and dishabituation by a novel stimulus, attributable to plasticity in the brain. Mutations of rutabaga that diminish cAMP synthesis reduced the rate of habituation, whereas dunce mutations that increase cAMP levels led to a detectable but moderate increase in habituation rates. Surprisingly, habituation was extremely rapid in dunce rutabaga double mutants. This corresponds to the extreme defects seen in double mutants in other learning tasks, and demonstrates that defects of the rutabaga and dunce products interact synergistically in ways that could not have been predicted on the basis of simple counterbalancing biochemical effects. Although habituation is localized to afferents to the giant fiber, cAMP mutations also affected performance of thoracic portions of the pathway on a millisecond time scale that did not account for behavioral plasticity. More significantly, spontaneous recovery and dishabituation were not as clearly affected as habituation in mutants, indicating that these processes may not overlap entirely in terms of cAMP-regulating mechanisms. The analysis of habituation of the giant fiber response in available learning and memory mutants could be a crucial step toward realizing the promise of memory mutations to elucidate mechanisms in neural circuits that underlie behavioral plasticity.

Altered mechanoreceptor response in Drosophila bang-sensitive mutants.

Bang-sensitive mutants of Drosophila melanogaster (bas1, bssMW1, eas2, tko25t) display seizure followed by paralysis when subjected to mechanical shock. However, no physiological or biochemical defect has been found to be common to all of these mutants. In order to observe the effects of bang-sensitive mutations upon an identified neuron, and to study the nature of mechanically induced paralysis, we examined the response of a mechanosensory neuron in these mutants. In each single mutant and the double mutant bas1 bssMW1, the frequency of action potentials in response to a bristle displacement was reduced. This is the first demonstration of a physiological defect common to several of the bang-sensitive mutations. Adaptation of spike frequency, cumulative adaptation to repeated stimulation (fatigue) and the time course of recovery from adaptation were also examined. Recovery from adaptation to a conditioning stimulus was examined in two mutants (bas1 and bssMW1), and initial recovery from adaptation was greater in both mutants. Quantification of receptor potentials was complicated by variability inherent in extracellular recording conditions, but examination of the waveform and range of amplitudes did not indicate clear mutant defects. Therefore the differences observed in the spike response may be due to an alteration of the transfer from receptor potentials to action potential production. DNA sequence analysis of tko and eas has indicated that they encode apparently unrelated biochemical products. Our results suggest that these biochemical lesions lead to a common physiological defect in mechanoreceptors.(ABSTRACT TRUNCATED AT 250 WORDS)

Interactions of membrane excitability mutations affecting potassium and sodium currents in the flight and giant fiber escape systems of Drosophila.

We have studied the influence of the K(+)-current mutations eag and Sh and the Na(+)-current mutation napts upon two well-defined neural circuits that underlie flight and an escape response in Drosophila, recording from dorsal longitudinal and tergotrochanteral muscles. Mutations of Sh and eag affected refractory period and following frequency, but not latency, of the jump-and-flight escape response. The napts mutation altered these 3 physiological parameters of the "jump" (TTM), but not the "flight" (DLM), branch, suggesting differences in the vulnerability of different circuit components to the mutation. In contrast to their interaction in some other systems, napts did not counteract the effects of eag and Sh upon these physiological parameters in eag Sh; nap triple mutants. In eag Sh double mutants, in which multiple K+ currents may be diminished, flight muscles showed abnormal rhythmic activity not associated with flight, and some flies also had an abnormal wings-down posture. The low-frequency spikes probably originated in the flight muscle motoneurons, but the coordination between muscle fibers during this "non-flight activity" was distinct from flight. Nevertheless, in spite of the presence of this non-flight activity in resting eag Sh flies, those animals with normal wing posture were also able to fly, with a normal pattern of muscle activity. This suggests that in these mutants, the DLM motoneuron circuit is able to switch between two patterns of output, non-flight activity and flight.(ABSTRACT TRUNCATED AT 250 WORDS)

Absolute configuration of glycerol derivatives. 4. Synthesis and pharmacological activity of chiral 2-alkylaminomethylbenzodioxans, competitive alpha-adrenergic antagonists.

The optical isomers of alpha-adrenergic receptor antagonists prosympal (2), piperoxan (3), and dibozane (4) were prepared by methods establishing the absolute configuration of each. (2S)-3(2'-Hydroxyphenoxy)-1,2-propanediol ditosylate (10) was prepared from (2R)-3-tosyloxy-1,2-propanediol acetonide (6). Intramolecular displacement afforded (2S)-tosyloxymethylbenzodioxan [(2R)-11]. Reaction of (2R)-11 with the appropriate amine (diethylamine, piperidine, or piperazine) afforded the 2S isomers of 2, 3, and 12, respectively. Reaction of (2S)-12 with (2R)-11 afforded the SS isomer of 4. Reaction of (2S)-3-benzyloxy-1,2-propanediol ditosylate (14) with catechol (NaOMe) afforded (2R)-benzyloxymethylbenzodioxan (15). Subjecting 15 to hydrogenolysis, tosylation, and displacement with the appropriate amine afforded 2R isomers of 2, 3, and 12. Reaction of (2R)-12 with (2S)-11 afforded (RR)-4. Reaction of (2R)-12 with (2R)-11 afforded meso-4. The S isomers were more effective antagonists to the alpha-adrenergic response of methoxamine-induced contraction of rabbit aortic strips by twofold in 2 and 18-19-fold in 3 and 4. meso-4 was as effective as the SS isomer of 4. The results are interpreted in terms of a similar conformational distribution of aminoalkyl, oxygen, and aromatic functional groups of the (S)-benzodioxans and (R)-epinephrine.