Voltage-Activated Currents From Adult Honeybee (Apis mellifera) Antennal Motor Neurons Recorded In Vitro and In Situ

Abstract

Voltage-activated currents from adult honey bee antennal motor neurons were characterized with in vitro studies in parallel with recordings taken from cells in situ. Two methods were used to ensure unequivocal identification of cells as antennal motor neurons: 1) selective backfilling of the neurons with fluorescent markers before dissociation for cell culture or before recording from cells in intact brains, semiintact brains, or in brain slices or 2) staining with a fluorescent marker via the patch pipette during recordings and identifying antennal motor neurons in situ on the basis of their characteristic morphology. Four voltage-activated currents were isolated in these antennal motor neurons with pharmacological, voltage, and ion substitution protocols. The neurons expressed at least two distinct K+ currents, a transient current (IA) that was blocked by 4-aminopyridine (4-5 x 10(-3) M), and a sustained current (IK(V)) that was partially blocked by tetraethylammonium (2-3 x 10(-2) M) and quinidine (5 x 10(-5) M). IA activated above -40 to -30 mV and the half-maximal voltages for steady-state activation and inactivation were -8.8 and -43.2 mV, respectively. IK(V) activated above -50 to -40 mV and the midpoint of the steady-state activation curve was +11.2 mV. IK(V) did not show steady-state inactivation. Additionally, two inward currents were isolated: a tetrodotoxin (10(-7) M)-sensitive, transient Na+ current (INa) that activated above -35 mV, with a maximum around -5 mV and a half-maximal voltage for inactivation of -72.6 mV, and a CdCl2 (5 x 10(-5) M)-sensitive Ca2+ current that activated above -45 to -40 mV, with a maximum around -15 mV. This study represents the first step in our effort to analyze the cellular and ionic mechanisms underlying the intrinsic properties and plasticity of antennal motor neurons.

Differential expression of voltage-sensitive K+ and Ca2+ currents in neurons of the honeybee olfactory pathway

“…Among the three different voltage-sensitive K + currents is a transient Atype K + current, which resembles the shaker-like current of other systems and interacts with a fast voltage-sensitive Na + current during spike generation (Pelz et al, 1999). Similar voltage-sensitive currents have been described in honeybee antennal motor neurons, both in vitro and in situ (Kloppenburg et al, 1999a).…”

Calcium Current Diversity in Physiologically Different Local Interneuron Types of the Antennal Lobe

“…In all investigated cell types, the current waveforms and physiological properties of I Ca were typical for voltage-activated Ca 2ϩ currents and in the range of other insect preparations [A. mellifera, antennal motorneurons (Kloppenburg et al, 1999a) and Kenyon cells (Schäfer et al, 1994); D. melanogaster, embryonic neurons (Byerly and Leung, 1988;Saito and Wu, 1991); Gryllus bimaculatus, giant interneurons (Kloppenburg and Hörner, 1998); P. americana, embryonic cockroach neurons (Benquet et al, 1999) and DUM neurons (Wicher and Penzlin, 1994); Manduca sexta, motor neurons (Hayashi and Levine, 1992); S. americana, thoracic neurons (Laurent et al, 1993); Schistocerca gregaria, DUM neurons (Heidel and Pflüger, 2006) and thoracic neurons (Pearson et al, 1993)]. However, a quantitative comparison of the functional properties from I Ca between the different cell types revealed significant differences in some physiologically important parameters (for summary, see supplemental Table 1, available at www.jneurosci.org as supplemental material).…”

“…5B). This is a relatively low activation threshold compared with I Ca in uPNs and in many other insect neurons (Byerly and Leung, 1988;Saito and Wu, 1991;Hayashi and Levine, 1992;Laurent et al, 1993;Pearson et al, 1993;Schä-fer et al, 1994;Wicher andPenzlin, 1994, 1997;Kloppenburg and Hörner, 1998;Benquet et al, 1999;Kloppenburg et al, 1999a;Heidel and Pflüger, 2006). In type II LNs, despite the similar activation threshold in both LN types, the half-maximal voltage for activation is significantly more hyperpolarized than in type I LNs (Fig.…”

“…4 D). This observation is remarkable because in most neuron types the voltage-activated Ca 2ϩ currents are masked by large outward K ϩ currents in absence of K ϩ channel blockers (Schäfer et al, 1994;Kloppenburg and Hörner, 1998;Benkenstein et al, 1999;Kloppenburg et al, 1999a). These Ca 2ϩ currents were of relatively large amplitude and are described in more detail below.…”

“…The solution was adjusted to pH 7.2 (with NaOH) and to 430 mOsm (with glucose). To isolate the Ca 2ϩ currents, a combination of pharmacological blockers and ion substitution was used that has been shown to be effective in central olfactory neurons of the cockroach (Husch et al, 2008) and in other insect preparations (Schäfer et al, 1994;Kloppenburg and Hörner, 1998;Kloppenburg et al, 1999a). Transient voltage-gated Na ϩ currents were blocked by tetrodotoxin (TTX) (10 Ϫ7 -10 Ϫ4 M; T-550; Alomone).…”