Naturally occuring polymorphisms in behavior are difficult to map genetically and thus are refractory to molecular characterization. An exception is the Drosophila melanogaster foraging (for) gene, which has two naturally occurring variants relating to food-search behavior: 'rover' and 'sitter'. Molecular mapping placed foraging mutations in the dg2 gene, which encodes a cyclic guanosine monophosphate (cGMP)-dependent protein kinase (PKG). Rovers have higher PKG activity than sitters, and transgenic sitters expressing a dg2 complementary DNA from rover show transformation of behavior to rover type. Thus, PKG levels affect food-search behavior, and natural variation in PKG activity accounts for a behavioral polymorphism (Osborne, 1997).

Interest in the cGMP-dependent serine/threonine kinase, or PKG, has grown with the awareness of the diversity of biochemical pathways that involve cGMP. PKG has been shown to influence characteristics involved in both functional and developmental plasticity of neural circuits. In Drosophila, one form of PKG (known as dg2; Kalderon, 1989) is encoded by the foraging gene (Osborne, 1997), which takes its name from a behavioral phenotype, the degree of locomotion while feeding, indicated by larval and adult foraging trail lengths (Sokolowski 1980; de Belle, 1987; de Belle, 1989; Pereira, 1993. Two naturally occurring variants, for^R (rovers, with long foraging trails) and for^s (sitters, with short foraging trails), have high and low PKG levels, respectively. Rovers and sitters do not differ in general activity in the absence of food. Both rovers and sitters are wild-type forms that exist at appreciable frequencies. Several mutations of the locus map with the naturally occurring alleles in the 24A3-5 region of the D. melanogaster polytene chromosomes. This region contains dg2, one of two cGMP-dependent protein kinase (PKG) genes in Drosophila. The dg2 gene has three major transcripts, T1, T2, and T3, and the for mutations are localized to this region. The P[GAL4] transposable element in 189Y was inserted in the 5' end of the dg2 T2 transcript. This homozygous viable insertion identified a new for allele, because P-element excision reverts larval foraging behavior from the sitter to the rover phenotype. As is the case with other sitter alleles, locomotion of the 189Y larvae is not reduced in the absence of food, indicating that the change in behavior is foraging-specific (Osborne, 1997).

To determine whether PKG is directly responsible for the foraging polymorphism in Drosophila, dg2 was overexpressed in sitter larvae. This results in a change of behavior to the rover phenotype. The transgenic strain contains four copies of a heat shock-driven dg2-cDNA. The basal level of PKG expression in this transgenic strain is sufficient to rescue rover larval behavior, thus eliminating the lethal and sublethal effects of heat on the dg2-transgenic larvae. As expected, the PKG enzyme activities of the dissected larval central nervous systems (CNSs) show that without heat shock, the dg2-cDNA transgenic strain have levels of PKG similar to those of for^R and significantly higher than those of the sitter control strain (Osborne, 1997).

The basis for the dg2 activity difference between for^R and for^s was further addressed by measurement of RNA levels and PKG protein. Northern (RNA) analysis revealed that for^s and for^s2 show a small but consistent reduction in the abundance of T1 RNA relative to that in for^R. T2 and T3 RNA are also reduced in these strains, but to a lesser extent. To assess protein levels, extracts of adult heads were subjected to protein immunoblot analysis by probing with an antibody to bovine PKG, or the extracts were affinity-purified by chromatography on cGMP-sepharose, labeled, and electrophoresed. In both experiments, a prominent band at a molecular mass of 80,000 Daltons was found. This is the only band strongly induced by heat shock in the dg2-cDNA transgenic strain, and it is less intense in for^s than for^R. (This band is also somewhat less intense in for^s2 and nearly absent in 189Y homozygotes). Taken together, these results argue that the difference between the naturally occurring alleles for^R and for^s is in the level of expression of the enzyme (Osborne, 1997).

The assignment of mutations in the for gene to the dg2 locus not only establishes the identification of PKG mutations but also implicates the cGMP signal transduction pathway in the regulation of food-search behavior in D. melanogaster. Small but significant differences in the levels of this kinase affect the naturally occurring behavioral polymorphism. These small differences in PKG are even detectable in homogenates, indicating that the differences in PKG level in rovers and sitters might be larger in cells relevant to the expression of the foraging behavior. These results suggest that the amount of kinase activity affects larval food-search behavior. Indeed, even modest quantitative changes in kinase activity affect behavior. Induced mutations that affect behavioral phenotypes often lie in signal transduction pathways. For example, the cyclic adenosine monophosphate (cAMP) system influences associative learning in flies, and genetic variants in two other serine/threonine kinases: the calcium/calmodulin-dependent protein kinase II and protein kinase C affect learning and behavioral plasticity in flies and mice. The finding that for encodes a PKG shows that a naturally occurring genetic polymorphism in behavior involves these pathways. PKG has a variety of pleiotropic cellular regulatory functions that are also typical of signal transduction components. Electrophysiological studies have shown that injected kinase affects neuronal membrane conductance in snails and mammals; that inhibitors of PKG block long-term potentiation in mammalian hippocampus and that PKG is involved in presynaptic long-term potentiation in cultured hippocampal neurons. Outside the nervous system, PKG has also been implicated in controlling proliferation of smooth muscle cells and neutrophil degranulation. These findings assign behavioral functions to this relatively scarce member of the serine/threonine kinases and show that subtle differences in PKG can lead to naturally occurring variation in behavior (Osborne, 1997 and references).

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. The electrically induced giant fiber response in intact tethered flies has been established as a model for habituation, a form of nonassociative learning. 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. Response decrement was assayed 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 is most rapid in genotypes previously shown to have low PKG activities and sitter-like foraging behavior. Differences were found 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 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 (Engel, 2000).

Habituation is a form of nonassociative learning in which a behavioral response is reduced or disappears with repeated stimulation. Nonassociative conditioning is of interest as a simple manifestation of physiological mechanisms that also may underlie more complex associative learning paradigms. Habituation may be mediated by a variety of mechanisms, including homosynaptic depression and extrinsic inhibition. Habituation is phylogenetically widespread and has functional significance in modulating both the gain and sensitivity of behavioral responses (Engel, 2000 and references therein).

The Drosophila giant fiber pathway that mediates the visually induced startle reflex, a jump-and-flight escape response, has been studied extensively at the levels of neural physiology and development. The response can be evoked by electrical stimulation to the brain in an intact animal, and this has allowed the bypassing of visual input and facilitated the focusing on central and motor stages of neural processing in an intact, behaviorally relevant circuit. The response likelihood diminishes with repeated electrical stimulation. This response decrement shows most of the typical characteristics of behavioral habituation including frequency dependence, strength dependence, habituation beyond zero response, spontaneous recovery, faster rehabituation, dishabituation, and habituation of dishabituation (Engel, 1996, 1998). Because electrical stimulation recruits the escape response circuit after initial stages of sensory processing, this report refers to modification patterns resembling 'habituation' and 'dishabituation' as 'response decrement' and 'evoked recovery,' respectively. Nevertheless, conformity to the characteristics of a widely studied learning paradigm makes the giant fiber response a useful model for genetic analyses of behavioral plasticity and its physiological correlates at the circuit level (Engel, 1996, 1998). This approach has provided evidence that Drosophila mutants defective in associative learning paradigms (in genes affecting cAMP metabolism) also display abnormal response decrement of the giant fiber response in a habituation protocol (Engel, 1996, 1998, 2000).

Several gene loci have been identified that influence habituation-like decrement of the giant fiber response, with products that include adenylyl cyclase (rutabaga) and cAMP phosphodiesterase (dunce; Engel, 1996), K+ channel subunits with distinct physiological properties including voltage activation (Shaker, ether à go-go), calcium activation (slowpoke), and channel modulation (Hyperkinetic; Engel, 1998), and now PKG (foraging). Like the cAMP pathway genes that affect learning, foraging has pleiotropic effects with potential fitness consequences (Hughes, 1996; Sokolowski, 1997; Wingrove, 1999). This pleiotropy is paralleled at the cellular level in which these gene products have diverse molecular targets and actions. PKG serine/threonine kinases have numerous targets that could affect neuronal function and growth, such as ion channels (Stockand, 1996; Carrier, 1997; Taguchi, 1997; Alioua, 1998; Han, 1998; Vaandrager, 1998; Wexler, 1998), ATPases (e.g., Uneyama, 1998), and regulators of gene expression (Gudi, 1997; Idriss, 1999). PKG may interact with other second messenger systems such as PKA, either by regulating such other systems (Moon, 1998) or by phosphorylating common targets (Lengyel, 1999). It is interesting that mutations of dunce that increase cAMP abundance lead to more rapid stimulus-dependent response decrement (Engel, 1996), opposite that of the effect of increased PKG activity in foraging rover genotypes (Engel, 2000).

Thus, a picture has emerged in which the molecular mechanisms that underlie response decrement in a habituation paradigm, like other neural plasticity such as LTP, are influenced by multiple biochemical and genetic factors. The redundancy of pathways influencing response modification therefore could allow habituation of the escape behavior to be modified and fine-tuned over the course of generations for more adaptive matching to ecologically relevant stimuli. An important point is that the foraging locus is known to be polymorphic in wild populations. This suggests that habituation of escape could vary among flies in a natural population. The foraging locus may be part of the genetic architecture through which plasticity and sensitivity of the escape response have been fine-tuned over evolutionary time (Engel, 2000).

The rate of response decrement has been found to be correlated with PKG activity and foraging behavior: decrement of the electrically induced response was most rapid in genotypes previously shown to have low PKG activity and sitter-like foraging behavior. Differences in spontaneous recovery from response decrement during a rest from stimulation and in dishabituation-like recovery were evoked by a novel stimulus (a puff of air). The data suggest that these differences in spontaneous recovery and evoked recovery may be secondary consequences of differing rates of response decrement. This indicates the interdependence of multiple processes of plasticity in stimulus-dependent response decrement of the giant fiber response. The data further raise the possibility that two processes with different time courses contribute to the response decrement (Engel, 2000).

Overall, the results show that PKG affects habituation-like response decrement in an identified neural circuit of intact tethered flies. From this it can be hypothesized that PKG also may be involved in other forms of learning. It has been shown that cAMP signaling pathways, which play an essential role in associative learning in flies, also affect stimulus-dependent decrement of the giant fiber response (Engel, 1996). The present results suggest that modulation of the escape response could involve the counterbalancing of multiple second messenger systems. A new adult-stage phenotype of the foraging locus has been defined. Finally, it has been shown that behaviorally relevant neural plasticity in an identified circuit can be influenced by a single-locus genetic polymorphism (Engel, 2000).

The response decrement of the long-latency giant fiber response induced has been examined by electrical stimulation; this treatment bypasses the initial stages of visual processing to recruit afferents to the descending giant fibers (Engel, 1996). Rates of response decrement are strongly affected by allelic variation in the foraging gene. Sitter stocks show more rapid response decrement than rovers in comparisons between the two artificially induced alleles or the two natural alleles. The most dramatic difference was between alleles generated artificially by P-element insertion and excision. for^189Y showed more rapid response decrement than any other line in this study. The abundance of foraging PKG is quite low in for^189Y (Osborne, 1997). In contrast, for^E1 showed scarcely any response decrement at the standard stimulation frequency of 5 Hz. In fact, some for^E1 flies could be driven at stimulus rates of 30 Hz or higher without showing failures. for^E1 arose by excision of the P-element from the foraging locus in for^189Y; rover behavior and high abundance of PKG are restored in for^E1 relative to for^189Y (Engel, 2000).

More subtle differences were observed between the naturally occurring alleles. for^s flies showed more rapid response decrement than for^R. Flies homozygous for each of the two foraging alleles for^R and for^s differ in their degrees of PKG activity (Osborne, 1997). for^R is genetically dominant to for^s for the larval foraging phenotype (de Belle, 1987) but intermediate for adult foraging (Pereira, 1993). As was the case for adult foraging behavior, heterozygous F₁ progeny (for^R/for^s) showed a rate of response decrement intermediate between the parental stocks, suggesting semidominance for this response modification phenotype (Engel, 2000).

The experiments described in this study were conducted within a single year (1999). The for^R and for^s stocks also were tested in this habituation-like protocol in 1996. In these earlier tests, the absolute resistance to response decrement was greater for both genotypes than in 1999, but for^s again showed more rapid response decrement than for^R. Similarly, repeated measurements of larval and adult foraging behavior have shown that it is the relative differences between rovers and sitters, not the absolute mean behavioral scores, that are maintained across tests performed at different times or in different laboratories (Engel, 2000).

Differences were observed in spontaneous recovery from decrement of the long-latency electrically induced response. Flies first were stimulated to a response decrement criterion of five consecutive failures (indicating a low response likelihood). One measure of spontaneous recovery is the response likelihood for the first stimulus given after 5 sec of rest. A 5-sec rest period is ordinarily sufficient for the response likelihood to return to nearly 100%, even in genotypes with very rapid response decrement (Engel, 1996, 1998). Full recovery of the response was observed for for^R and for^s flies as well as for^R/for^s heterozygotes. However, for^189Y flies did not recover fully in 5 sec (Engel, 2000).

A second measure of recovery is the resistance to response decrement within a subsequent stimulus episode. This was quantified as the number of responses evoked before reaching the five-failure decrement criterion during stimulus bouts delivered after different recovery intervals. The resistance to response decrement integrates performance over the entire stimulus bout rather than just the first stimulus. The for genotypes showed clear differences in their degree of recovery to initial rates of response decrement. Absolute postrecovery response numbers were highest for for^R, the most slowly decrementing stock, and were progressively lower for more rapidly decrementing genotypes. However, when mean response numbers were divided by first-bout response numbers to give normalized recovery indices, this ranking was reversed: the highest recovery indices were shown by rapidly decrementing sitter genotypes, particularly after 30- and 120-sec recovery intervals. When postrecovery response scores were log-transformed, effectively normalizing the results within genotypes while retaining scale differences between genotypes, the kinetic profiles of recovery showed a similar ranking pattern, with the greatest degrees of recovery after 30- and 120-sec intervals being shown by rapidly decrementing genotypes (Engel, 2000).

The slight degree of spontaneous recovery between 30 and 120 sec suggests that, in addition to a short-term component of response decrement that recovers in less than 30 sec, there is also a long-term component of response decrement with slower onset and recovery kinetics that becomes stronger over multiple stimulus bouts and recovers with a time course exceeding 120 sec. In previous work, 30- or 120-sec recovery intervals were tested after a single prior stimulus bout (in different groups of flies), and with that protocol the recovery to first-bout response decrement rates was nearly complete (Engel, 1996, 1998). In the present experiments, each fly received four stimulus bouts separated by intervals of 5, 30, and 120 sec, so that 30- and 120-sec recovery intervals were tested after two or three prior stimulus bouts (instead of one prior bout as in the earlier studies). It appears that additional prior stimulus bouts affected the state of the response pathway even though every bout ended with a consistent response decrement criterion of five failures (Engel, 2000).

A slowly developing component of response decrement could be most apparent in slowly decrementing flies, because they are exposed to a greater number of stimuli in the two or three bouts preceding the recovery interval. Consistent with this, the lowest 30- and 120-sec recovery indices were shown by the most slowly decrementing genotypes. To examine this relationship more directly, normalized recovery indices for individual flies of all genotypes were plotted against the total number of stimuli given in bouts before the recovery interval. After 30- or 120-sec intervals, recovery indices were inversely related to the number of prior stimuli. This relationship was most evident for the range of 50 to 300 prior stimuli, suggesting that this slow component of response decrement became saturated after 300 stimuli and that other factors contributed more to response variation with fewer than 50 prior stimuli. After the shortest recovery interval of 5 sec, the relationship was weak. This suggests that recovery from a short-term process of response decrement is the predominant factor during the first 5 sec after the end of a bout (Engel, 2000).

The potential to distinguish multiple components of habituation-like response decrement in this system will require further study. Here, it is most important to note that for genotypes showed differences in recovery when tested under a consistent protocol (Engel, 2000).

Recovery of the long-latency giant fiber response can be evoked by a novel stimulus such as an airpuff in a dishabituation protocol (Engel, 1996, 1998). Clear evoked recovery could be shown in each strain except for^189Y. The number of responses for the 20 stimuli after an airpuff or 'sham puff' (each averaged from five repetitions) was divided by the number of responses at the beginnings of bouts, giving test and control scores, respectively. The operational criterion for evoked recovery was a test score greater than double the control score. Evoked recovery was observed most often in slowly decrementing genotypes (for^R and for^R/for^s). Among flies that did show evoked recovery by this definition, the magnitude of recovery (the test score) was also greatest in slowly decrementing genotypes (Engel, 2000).

Few for^E1 flies showed response decrement to five-failure criterion at the standard stimulation frequency of 5 Hz. However, with higher stimulus frequencies for^E1 flies did display habituation-like response decrement, characterized by synchronous loss of responses in DLM (Dorsal Longitudinal Muscle) and TTM (Tergotrochanteral Muscle), spontaneous recovery, and recovery evoked by an airpuff (Engel, 2000).

Latency and refractory period are indicators of the integrity of neural connectivity and signal transmission in the giant fiber pathway. Two response classes, evoked by different stimulus voltages, give information about different parts of the circuit. Weak stimuli evoke a long-latency response by recruiting afferent neurons upstream of the giant fibers, whereas stronger stimuli trigger a short-latency response by directly activating the giant fibers (Engel, 1996). The long-latency response can reveal properties of connections in the brain that do not contribute to the short-latency response. The thoracic portion of the circuit (activated in both long- and short-latency responses) can give information about how mutations affect neural functioning within a network of identified neurons. The TTM branch has a single electrochemical neuronal synapse onto the TTM motoneuron, whereas the DLM branch includes two synapses, an apparent electrochemical synapse of the cervical giant fiber onto the peripherally synapsing interneuron (PSI) neuron and cholinergic synapses of the PSI onto the DLM motoneurons (Engel, 2000 and references therein).

Response latencies differed between for^E1 and for^189Y for the long-latency response but not the short-latency one. Latency (but not refractory period or response decrement in a habituation protocol) is significantly influenced by ambient temperature (Engel, 1996). The response latencies was tested for for^E1 and for^189Y under similar temperature conditions and during the same period of days. Response latencies did not differ when other genotypes were compared (Engel, 2000).

The twin-pulse refractory period of the short-latency response, mediated in the thoracic portion of the giant fiber pathway, has proven to be a sensitive indicator of deficits in basic physiological properties such as transmitter processing and ion channel function. Short-latency response refractory periods were not significantly affected by allelic variation at the foraging locus. The refractory period of the long-latency response, mediated in the afferent portion of the pathway, is an indicator of properties of the brain portion of the circuit (Engel, 1996, 1998). The long-latency refractory period tended to be shorter in genotypes with slower stimulus-dependent response decrement. This is most clear when for^E1 and for^189Y are compared (Engel, 2000).

It is interesting that for^E1 and for^189Y showed differences in response properties that were restricted to the afferent portion of the neural pathway, because these stocks showed an extreme difference in response decrement in the habituation protocol, which also is mediated in the afferent portion of the pathway. Despite these differences, it is clear that the giant fiber pathway is fundamentally sound in all the foraging genotypes tested. The extreme effects on response latency or short-latency refractory period that have been reported using mutations affecting ion channels or synaptic integrity were not found in genotypes differing in PKG activity (Engel, 2000).

These results strongly indicate that the foraging PKG affects habituation-like response decrement in the electrically induced giant fiber response. Artificially induced alleles (for^E1 and for^189Y) defined the influence of PKG in response decrement of the giant fiber response, and more modest naturally occurring genetic variants (for^R and for^s) showed similar but more subtle effects. In comparisons between different genotypes at the PKG foraging locus, response decrement was slower in genotypes with more abundant PKG (for^E1 and for^R) than in genotypes with less abundant PKG (for^189Y and for^s). It is interesting that rate of response decrement, response latency, and refractory period were all more extreme in for^E1 than the wild rover genotype for^R. It is possible that imprecise excision of the P-element from for^189Y resulted in a more highly expressing allele in for^E1 than the original parental for allele from which for^189Y arose. Sequencing of for^E1, currently in progress, should help to resolve this possibility. Differences in rate of response decrement followed a semidominant mode of inheritance as shown by for^R/for^s heterozygotes. Semidominant inheritance also has been reported (Pereira, 1993) for the adult rover and sitter foraging phenotypes (Engel, 2000).

Recovery results indicate that foraging affects spontaneous recovery from stimulus-dependent response decrement. The results also imply the existence of distinct components of this habituation-like response decrement with different kinetics of onset and recovery that could partly account for genetic differences in recovery phenotypes. A long-term component of response decrement is suggested by the similarity of recovery indices after either 30- or 120-sec recovery intervals. For those intervals, recovery of the resistance to subsequent response decrement is correlated with the number of stimuli that were given before the recovery rest interval (Engel, 2000).

Sitter genotypes with low PKG expression showed the greatest recovery of resistance to response decrement after 30- and 120-sec intervals. However, these flies also showed more rapid response decrement in initial stimulus bouts and experienced fewer stimuli in all bouts before recovery testing, and in consequence may have had less exposure to a long-term component of response decrement. Therefore, differences in rates of response decrement may have contributed indirectly to the observed genetic differences in recovery indices for 30 and 120 sec. This would not preclude the possibility that PKG also could play a role in physiological processes that underlie spontaneous recovery per se (Engel, 2000).

Early recovery after stimulus-dependent response decrement appears to be dominated by a short-term component of response decrement. Recovery indices increased substantially between 5 and 30 sec after ending the preceding stimulus bout, and response likelihood did not recover to 100% after 5 sec in some genotypes. Response likelihood for the first stimulus following a 5-sec recovery interval showed complete recovery in for^s and for^R but did not recover completely in for^189Y flies, which showed the most rapid response decrement in this study and have low PKG expression (Osborne, 1997). In contrast to the recovery of resistance to subsequent response decrement, this genetic effect could not be a consequence of differences in exposure to a long-term response decrement process, because for^189Y flies actually experienced the smallest numbers of stimuli before the 5-sec recovery interval. This result suggests that PKG may facilitate recovery of the likelihood of responding to a single stimulus after prior response decrement (Engel, 2000).

Evoked recovery in a dishabituation protocol was weakest in the most rapidly decrementing foraging genotypes. These results may point to a direct involvement of PKG pathways in evoked recovery. Alternatively, a more rapid rate of response decrement in sitter genotypes could have reduced evoked recovery in an indirect manner as follows. Assuming an equivalent activation of recovery processes by an airpuff in all genotypes, more rapid response decrement after the puff could diminish the amount of recovery observed. Furthermore, because a standard decrement criterion of five consecutive failures preceded the puff in all genotypes, a rapid rate of 'latent' response decrement during the five criterion stimuli could induce a deeper level of response decrement for the circuit to recover from at the time of the airpuff (Engel, 2000).

These results suggest that the foraging PKG could affect the observed levels of spontaneous recovery and evoked recovery in part through altering the rate of stimulus-dependent response decrement. Similar correspondences between response decrement rates, spontaneous recovery, and evoked recovery may be seen for cAMP metabolic mutants. This highlights the interrelatedness of these three processes in the giant fiber system. One goal for the future is to determine the extent to which these phenomena can be altered independently by mutations and thus may involve independent molecular mechanisms (Engel, 2000).

Differences were seen in the response latencies and refractory periods of the for^E1 and for^189Y genotypes. These effects were seen in the long-latency response but not the short-latency response, indicating that they are mediated in the afferent or brain segment of the giant fiber pathway in which habituation-like response decrement also is mediated (Engel, 1996, 1998). However, response decrement rate and long-latency refractory period may not be functionally related. Earlier studies with mutations affecting cAMP cascades (Engel, 1996) and K₊ channels (Engel, 1998) have not shown a strong correlation between response decrement in a habituation protocol and refractory period. Moreover, flies bearing Shaker and ether à go-go K₊ channel mutations have refractory periods comparable to for^E1 but show much more rapid response decrement (Engel, 1998). Previous studies (Engel, 1992, 1996, 1998) have indicated that the thoracic portion of the giant fiber pathway may have qualitatively normal characteristics even in genotypes with very rapid response decrement. Consistent with this, short-latency refractory periods and latencies did not differ between foraging genotypes, even the very rapidly decrementing genotype for^189Y (Engel, 2000).

The results associate high PKG expression with a slow rate of response decrement in a habituation protocol but do not indicate the mechanism underlying this association. PKG may play a direct role in plasticity, either by down-regulating a physiological process that underlies response decrement or by enhancing a concomitant process of response sensitization as in a dual process model. Alternatively, high PKG expression could influence response decrement in a less direct manner by modifying the physiological or developmental context in which it occurs. For instance, if PKG enhanced basic properties of neural conduction or synaptic transmission so that the neural signal is stronger to begin with, then it could take longer for normally functioning mechanisms underlying stimulus-dependent response decrement to lead to failed responses. Enhancement of neural response properties would be consistent with the for^E1 phenotype of shortened latency and refractory period of the long-latency response, parameters that are mediated in the afferent part of the giant fiber pathway, just as is the habituation-like response decrement is (Engel, 2000).

PKG appears to affect such basic functional properties differently in different parts of the fly nervous system. Variation in foraging genotype did not affect latency or refractory period of the short-latency response, parameters that are mediated in the thoracic portion of the pathway. Moreover, reduced PKG activity in sitter genotypes is associated with hyperexcitability and enhanced nerve terminal sprouting at larval neuromuscular junctions and with reduced K₊ currents and increased membrane excitability in a significant population of neurons in dissociated embryonic cultures (Renger, 1999). These observations suggest a widely distributed role for PKG in the nervous system of flies (Engel, 2000).

GENE STRUCTURE

cDNA clone length - 4105bp (for transcript A - there are additional transcripts)

Bases in 5' UTR - 776

Exons - 9

Bases in 3' UTR - 644

PROTEIN STRUCTURE

Amino Acids - 894 (plus additional polypeptides)

Structural Domains

Two Drosophila genes encoding products related to cGMP-dependent protein kinase have been isolated by cross-hybridization to a Drosophila cAMP-dependent protein kinase catalytic subunit gene. Both genes encode products with putative cGMP binding and kinase domains on the same polypeptide chain, as found for the prototypical bovine lung cGMP-dependent protein kinase. The deduced product of one gene (DG1; cytological position, 21D) is 14% larger than the bovine enzyme and differs substantially in sequence at the amino terminus, the region responsible in the bovine enzyme for dimerization. The second gene (DG2; cytological position, 24A) is transcribed into three major RNA species of different size. The largest (DG2; T1) and smallest (DG2;T3) RNAs encode overlapping polypeptides of similar sequence to the whole length of bovine lung cGMP-dependent protein kinase. The translation product of the third major RNA (DG2;T2) lacks sequences similar to those that constitute the dimerization and kinase inhibitory domains of the bovine enzyme. The percentage amino acid identity between DG1 or DG2 and bovine lung cGMP-dependent protein kinase is 55% and 64%, respectively. A common progenitor of the two cGMP-dependent protein kinase genes, DG1 and DG2, is strongly suggested by the conserved positions of introns in these genes (Kalderon, 1989).

date revised: 12 September 2004

Home page: The Interactive Fly © 1995, 1996 Thomas B. Brody, Ph.D.

The Interactive Fly resides on the
Society for Developmental Biology's Web server.