Brian Waldrop

Local interneurons and information processing in the olfactory glomeruli of the moth Manduca sexta

Intracellular recordings were made from the major neurites of local interneurons in the moth antennal lobe. Antennal nerve stimulation evoked 3 patterns of postsynaptic activity: (i) a short-latency compound excitatory postsynaptic potential that, based on electrical stimulation of the antennal nerve and stimulation of the antenna with odors, represents a monosynaptic input from olfactory afferent axons (71 out of 86 neurons), (ii) a delayed activation of firing in response to both electrical- and odor-driven input (11 neurons), and (iii) a delayed membrane hyperpolarization in response to antennal nerve input (4 neurons).Simultaneous intracellular recordings from a local interneuron with short-latency responses and a projection (output) neuron revealed unidirectional synaptic interactions between these two cell types. In 20% of the 30 pairs studied, spontaneous and current-induced spiking activity in a local interneuron correlated with hyperpolarization and suppression of firing in a projection neuron. No evidence for recurrent or feedback inhibition of projection neurons was found. Furthermore, suppression of firing in an inhibitory local interneuron led to an increase in firing in the normally quiescent projection neuron, suggesting that a disinhibitory pathway may mediate excitation in projection neurons. This is the first direct evidence of an inhibitory role for local interneurons in olfactory information processing in insects. Through different types of multisynaptic interactions with projection neurons, local interneurons help to generate and shape the output from olfactory glomeruli in the antennal lobe.

GABA-mediated synaptic inhibition of projection neurons in the antennal lobes of the sphinx moth, Manduca sexta

Summary1.Responses of neurons in the antennal lobe (AL) of the mothManduca sexta to stimulation of the ipsilateral antenna by odors consist of excitatory and inhibitory synaptic potentials (Fig. 2A). Stimulation of primary afferent fibers by electrical shock of the antennal nerve causes a characteristic IPSP-EPSP synaptic response in AL projection neurons (Fig. 2B).2.The IPSP in projection neurons reverses below the resting potential (Fig. 3), is sensitive to changes in external (Fig. 4) and internal (Fig. 5) chloride concentration, and thus is apparently mediated by an increase in chloride conductance.3.The IPSP is reversibly blocked by 100 μM picrotoxin (Fig. 6) or bicuculline (Fig. 7).4.Many AL neurons respond to application of GABA with a strong hyperpolarization and an inhibition of spontaneous spiking activity (Fig. 8). GABA responses are associated with an increase in neuronal input conductance (Fig. 9) and a reversal potential below the resting potential (Fig. 11).5.Application of GABA blocks inhibitory synaptic inputs (Fig. 12 A) and reduces or blocks excitatory inputs (Fig. 12B). EPSPs can be protected from depression by application of GABA (Fig. 12B).6.Muscimol, a GABA analog that mimics GABA responses at GABAA receptors but not at GABAB receptors in the vertebrate CNS, inhibits many AL neurons in the moth (Fig. 13).

Multitasking in the Olfactory System: Context-Dependent Responses to Odors Reveal Dual GABA-Regulated Coding Mechanisms in Single Olfactory Projection Neurons

Studies of olfaction have focused mainly on neural processing of information about the chemistry of odors, but olfactory stimuli have other properties that also affect central responses and thus influence behavior. In moths, continuous and intermittent stimulation with the same odor evokes two distinct flight behaviors, but the neural basis of this differential response is unknown. Here we show that certain projection neurons (PNs) in the primary olfactory center in the brain give context-dependent responses to a specific odor blend, and these responses are shaped in several ways by a bicuculline-sensitive GABA receptor. Pharmacological dissection of PN responses reveals that bicuculline blocks GABAA-type receptors/chloride channels in PNs, and that these receptors play a critical role in shaping the responses of these glomerular output neurons. The firing patterns of PNs are not odor-specific but are strongly modulated by the temporal pattern of the odor stimulus. Brief repetitive odor pulses evoke fast inhibitory potentials, followed by discrete bursts of action potentials that are phase-locked to the pulses. In contrast, the response to a single prolonged stimulus with the same odor is a series of slow oscillations underlying irregular firing. Bicuculline disrupts the timing of both types of responses, suggesting that GABAA-like receptors underlie both coding mechanisms. These results suggest that glomerular output neurons could use more than one coding scheme to represent a single olfactory stimulus. Moreover, these context-dependent odor responses encode information about both the chemical composition and the temporal pattern of the odor signal. Together with behavioral evidence, these findings suggest that context-dependent odor responses evoke different perceptions in the brain that provide the animal with important information about the spatiotemporal variations that occur in natural odor plumes.

Physiology and pharmacology of acetylcholinergic responses of interneurons in the antennal lobes of the mothManduca sexta

Summary1.Neurons in the antennal lobe (AL) of the mothManduca sexta respond to the application, via pressure injection into the neuropil, of acetylcholine (ACh). When synaptic transmission is not blocked, both excitatory (Fig. 2) and inhibitory (Fig. 3) responses are seen.2.Responses to ACh appear to be receptor-mediated, as they are associated with an increase in input conductance (Figs. 2B and 3B) and are dose-dependent (Fig. 2C).3.All neurons responsive to ACh are also excited by nicotine. Responses to nicotine are stronger and more prolonged than responses to ACh (Fig. 4C). No responses are observed to the muscarinic agonist, oxotremorine (Fig. 4B).4.Curare blocks responses of AL neurons to applied ACh, while atropine and dexetimide are only weakly effective at reducing ACh responses (Figs. 5 and 6).5.Curare is also more effective than atropine or dexetimide at reducing synaptically-mediated responses of AL neurons (Fig. 7).6.In one AL neuron, bicuculline methiodide (BMI) blocked the IPSP produced by electrical stimulation of the antennal nerve, but it did not reduce the inhibitory response to application of ACh (Fig. 8).

Development of the gin trap reflex inManduca sexta: a comparison of larval and pupal motor responses

Summary1.Responses of motor neurons in larvae and pupae ofManduca sexta to stimulation of tactile sensory neurons were measured in both semiintact, and isolated nerve cord preparations. These motor neurons innervate abdominal intersegmental muscles which are involved in the production of a general flexion reflex in the larva, and the closure reflex of the pupal gin traps.2.Larval motor neurons respond to stimulation of sensory neurons innervating abdominal mechanosensory hairs with prolonged, tonic excitation ipsilaterally, and either weak excitation or inhibition contralaterally (Figs. 4A, 6).3.Pupae respond to tactile stimulation of mechanosensory hairs within the gin traps with a rapid closure reflex. Motor neurons which innervate muscles ipsilateral to the stimulus exhibit a large depolarization, high frequency firing, and abrupt termination (Figs. 2, 4B). Generally, contralateral motor neurons fire antiphasically to the ipsilateral motor neurons, producing a characteristic triphasic firing pattern (Figs. 7, 8) which is not seen in the larva.4.Pupal motor neurons can also respond to sensory stimulation with other types of patterns, including rotational responses (Fig. 3 A), gin trap opening reflexes (Fig. 3 B), and ‘flip-flop’ responses (Fig. 9).5.Pupal motor neurons, like larval motor neurons, do not show oscillatory responses to tonic current injection, nor do motor neurons of either stage appear to interact synaptically with one another. Most pupal motor neurons also exhibit i-V properties similar to those of larval motor neurons (Table 1; Fig. 10). Some pupal motor neurons, however, show a marked non-linear response to depolarizing current injection (Fig. 11).

Intersegmental interneurons serving larval and pupal mechanosensory reflexes in the moth Manduca sexta

Summary1.Intersegmental interneurons (INs) that participate in the larval bending reflex and the pupal gin trap closure reflex were identified in the isolated ventral nerve cord of Manduca sexta. INs 305, 504, and 703 show qualitatively different responses in the pupa than in the larva to electrical stimulation of sensory neurons that are retained during the larval-pupal transition to serve both reflexes. Action potentials produced by current injected into the 3 interneurons excite motor neurons that are directly involved in the larval and pupal reflexes. The excitation of the motor neurons is not associated with EPSPs at a fixed latency following action potentials in the interneurons, and thus there do not seem to be direct synaptic connections between the interneurons and the motor neurons.2.IN 305 (Fig. 2) has a lateral soma, processes in most of the dorsal neuropil ipsilateral to the soma, and a crossing neurite that gives rise to a single contralateral descending axon. IN 305 is excited by stimulation of the sensory nerve ipsilateral to its soma in the larva and the pupa. Stimulation of the sensory nerve contralateral to its soma produces an inhibitory response in the larva, but a mixed excitatory/inhibitory response to the identical stimulus in the pupa.3.IN 504 (Fig. 3) has a lateral soma, processes throughout most of the neuropil ipsilateral to the soma, and a crossing neurite that bifurcates to give rise to a process extending to the caudal limit of the neuropil and an ascending axon. IN 504 is excited by stimulation of the sensory nerve ipsilateral to its soma in both larvae and pupae, while the response to stimulation of the sensory nerve contralateral to its soma is inhibitory in the larva but mixed (excitatory/inhibitory) in the pupa.4.IN 703 has a large antero-lateral soma, a neurite that extends across to the contralateral side giving rise to processes located primarily dorsally in both ipsilateral and contralateral neuropils, and two axons that ascend and descend in the connectives contralateral to the soma (Fig. 4). IN 703 responds to stimulation of the sensory nerves on either side of the ganglion, but the form of the response changes during the larval-pupal transition. In the larva, the response consists of very phasic (0–2 spikes) excitation, but in the pupa there is a prolonged excitation that greatly outlasts the stimulus (Fig. 6).5.While the resting potential, and thus the relative spike threshold, of IN 703 appears to change during the larval-pupal transition (Fig. 9), hyperpolarizing IN 703 during a response shows that this difference can not account for the change in response properties (Fig. 10). Rather, IN 703 in the pupa is influenced by interneuronal inputs in the pupa whose effects are not expressed in the larva (Fig. 11).

Neurotransmitters and Neuropeptides in the Olfactory Pathway of the Sphinx Moth Manduca Sexta

Like other animals, insects have many and diverse chemical “messengers” in their nervous systems. A growing list of synaptic neurotransmitters, neuromodulators, neuropeptides, and neurohormones -- collectively “transmitters” -- prompts efforts to seek physiological roles and mechanisms of action for these substances. An improved understanding of these chemical messengers in the insect nervous system promises to reveal key regulatory mechanisms, novel and accessible targets for pharmacological agents, and phyletic differences that can be exploited in new approaches to the manipulation of insect behavior and the selective destruction of harmful populations of insect pests and disease vectors. Toward such goals, we study the biochemistry, cellular distribution, and physiological actions of transmitter candidates in an experimentally favorable insect “model”, the sphinx moth Manduca sexta. In contrast with significant advances that have been made in many laboratories investigating peripheral neural and neuromuscular systems, relatively much less is known about chemical signalling in the insect central nervous system (CNS). With this in mind, and building upon substantial previous and ongoing studies of the anatomy, physiology, and development of the insect CNS in many laboratories including our own, we focus on the cellular neurochemistry of the CNS in Manduca. In particular we are exploring the olfactory pathway in the brain, for which we have accumulated much information about the types of neurons and their functional organization and development (e.g. see recent reviews: Hildebrand, 1985; Hildebrand and Montague, 1986; Christensen and Hildebrand, 1987).