Leucokinin VIII

An antiserum against an achetakinin analog selectively localized leucokinin VIII (LKVIII) in the CNS of Leucophaea maderae. Preabsorption studies of the achetakinin antiserum with either preimmune serum or LKVIII prevented a positive reaction in both ELISA and immunocytochemical procedures. LKVIII immunoreactive neurons were found in the brain, frontal, and subesophageal ganglion, all 3 thoracic ganglia and the terminal ganglion. Nerves originating from the thoracic and terminal abdominal ganglia contain LKVIII material. Lateral and medial neurosecretory cells synthesizing LKVIII-like products contribute axons to the nervi corporis cardiaci that terminate in neurohemal sites in the corpora cardiaca and nervi corporis allati. Thus, leucokinin VIII, like leucokinin I (LKI) and leucomyosuppressin (LMS), appears to have both a neurohemal and neurotransmitter mode of regulating target cells in L. maderae.

The diuretic peptide leucokinin is known to increase fluid secretion in Malpighian tubules of the yellow fever mosquito Aedes aegypti by increasing a transepithelial Cl(-) conductance. The present study sought to examine whether stellate cells provided this transepithelial conductance in Aedes Malpighian tubules as they do in Drosophila Malpighian tubules. Aedes Malpighian tubule segments with and without stellate cells were perfused in vitro for measurements of the transepithelial voltage (V(t)), resistance (R(t)) and Cl(-) diffusion potentials (DP(Cl)). In 11 tubule segments containing both principal cells and stellate cells, 1 micro mol l(-1) leucokinin-VIII added to the peritubular bath immediately and significantly decreased V(t) from 39.3+/-14.3 mV to 2.3+/-0.7 mV, decreased R(t) from 12.4+/-2.6 kOmegacm to 2.4+/-0.3 kOmegacm, and increased DP(Cl) from 8.2+/-1.2 mV to 42.1+/-5.4 mV. These effects of leucokinin-VIII were qualitatively and quantitatively similar in six tubule segments containing no stellate cells; V(t) decreased from 37.8+/-7.0 mV to 3.4+/-0.6 mV, R(t) decreased from 8.8+/-2.1 kOmegacm to 1.7+/-0.2 kOmegacm, and DP(Cl) increased from 5.8+/-2.6 mV to 50.0+/-2.1 mV. Thus, stellate cells are not required for signaling or mediating the effects of leucokinin in Malpighian tubules of Aedes aegypti. The results further support previous observations that principal cells signal the effects of leucokinin to increase the Cl(-) conductance of the paracellular pathway through septate (or tight) junctions.

The role of Ca(2+) in mediating the diuretic effects of leucokinin-VIII was studied in isolated perfused Malpighian tubules of the yellow fever mosquito, Aedes aegypti. Peritubular leucokinin-VIII (1 microM) decreased the transepithelial resistance from 11.2 to 2.6 kOmega. cm, lowered the transepithelial voltage from 42.8 to 2.7 mV, and increased transepithelial Cl(-) diffusion potentials 5.1-fold. In principal cells of the tubules, leucokinin-VIII decreased the fractional resistance of the basolateral membrane from 0.733 to 0.518. These effects were reversed by the peritubular Ca(2+)-channel blocker nifedipine, suggesting a role of peritubular Ca(2+) and basolateral Ca(2+) channels in signal transduction. In Ca(2+)-free Ringer bath, the effects of leucokinin-VIII were partial and transient but were fully restored after the bath Ca(2+) concentration was restored. Increasing intracellular Ca(2+) with thapsigargin duplicated the effects of leucokinin-VIII, provided that peritubular Ca(2+) was present. The kinetics of the effects of leucokinin-VIII is faster than that of thapsigargin, suggesting the activation of inositol-1,4,5-trisphosphate-receptor channels of intracellular stores. Store depletion may then bring about Ca(2+) entry into principal cells via nifedipine-sensitive Ca(2+) channels in the basolateral membrane.