Motor pattern selection via inhibition of parallel pathways

Abstract

Motor pattern selection from a multifunctional neural network often results from direct synaptic and modulatory actions of different projection neurons onto neural network components. Less well documented is the presence and function of interactions among distinct projection neurons innervating the same network. In the stomatogastric nervous system of the crab Cancer borealis, several distinct projection neurons that influence the pyloric and gastric mill rhythms have been studied. These rhythms are generated by overlapping subsets of identified neurons in the stomatogastric ganglion (STG). One of these identified projection neurons is the modulatory proctolin neuron (MPN). We showed previously that MPN stimulation excites the pyloric rhythm by its excitatory actions on STG neurons. In contrast to its excitatory actions on the pyloric rhythm, we have now found that MPN inhibits the gastric mill rhythm. This inhibition does not occur within the STG, but instead results from MPN-mediated inhibition of two previously identified projection neurons within the commissural ganglia. These projection neurons innervate the STG and, via their actions on STG neurons, they elicit the gastric mill rhythm as well as modify the pyloric rhythm in a manner distinct from MPN. By inhibiting these projection neurons, MPN removes excitatory drive to gastric mill neurons and elicits an MPN-specific pyloric rhythm. Motor pattern selection by MPN therefore results from both a direct modulation of STG network activity and an inhibition of competing pathways.

Fig. 1.

Schematics of the STNS, including somata location and axonal pathway of the identified projection neurons MPN, MCN1, and CPN2. A, There is a pair of MPN somata located either in the OG or in the nerve posterior to this ganglion. Each MPN projects an axon through each superior esophageal nerve (son) to the CoG and through the stomatogastric nerve (stn) to the STG. It also projects an axonal branch from each superior esophageal nerve through the peripheral nerve dorsal posterior esophageal nerve (dpon). For clarity, the complete projection of only one MPN is shown. B, There is a single MCN1 and CPN2 in each CoG. Each MCN1 projects through the inferior esophageal nerve (ion) and stomatogastric nerve to the STG. Each CPN2 projects through the superior esophageal nerve and stomatogastric nerve to the STG. For clarity, the complete projection of only one MCN1 and one CPN2 is shown. CoG, Commissural ganglion;OG, esophageal ganglion; dgn, dorsal gastric nerve; lgn, lateral gastric nerve;lvn, lateral ventricular nerve; mvn, medial ventricular nerve; pdn, pyloric dilator nerve;CPN2, commissural projection neuron 2;MCN1, modulatory commissural neuron 1;MPN, modulatory proctolin neuron. Anterior is toward thetop of the figure and posterior is toward thebottom.

Fig. 2.

Examples of gastric mill rhythms elicited by MCN1 alone and by conjoint activity in MCN1 and CPN2. A,Left, Extracellular recordings of STG motor nerves during an MCN1-elicited gastric mill rhythm (mvn,dgn, lgn) and pyloric rhythm (pdn). This gastric mill rhythm is characterized by rhythmic alternating bursting in the LG (lgn) and DG (dgn) neurons. The VD neuron (smaller unit in themvn) is silent during each LG burst, and IC (larger unit in the mvn) is silent during each DG burst.Right, Schematic of the time of activity of STG neurons during an MCN1-elicited gastric mill rhythm. This gastric mill rhythm was elicited by selective stimulation of MCN1 (see also Coleman and Nusbaum, 1994; Coleman et al., 1995). B,Left, Extracellular recordings monitoring the pyloric and gastric mill rhythms during coactivation of MCN1 and CPN2. This gastric mill rhythm differs in several ways from the rhythm elicited by selective activation of MCN1. The cycle period is longer, both IC and VD (mvn) are completely inhibited during each LG burst, and the GM neurons (smallest unit in dgn) are strongly activated during each LG burst (see also Norris et al., 1994a).Right, Schematic of the time of activity of STG neurons during an MCN1/CPN2-elicited gastric mill rhythm. In this recording, MCN1 and CPN2 were activated by bath application of 10⁻⁵m oxotremorine to the entire preparation. The tonically active unit in the dgn in these and all subsequent dgn recordings is the anterior gastric receptor sensory neuron. Dark cell bodies represent active neurons; light cell bodies represent inactive neurons.

Fig. 3.

MPN activity inhibits the gastric mill rhythm.A, An ongoing MCN1-elicited gastric mill rhythm is suppressed by intracellular stimulation of MPN (firing frequency, 13 Hz). This resulted in termination of activity in the LG and DG neurons and the elimination of gastric mill-timed inhibition of the VD and IC neurons. The gastric mill rhythm resumed 10 sec after the end of the MPN stimulation. B, An MCN1/CPN2-elicited gastric mill rhythm was evoked by bath application of 10⁻⁶m oxotremorine and 10⁻⁷mF₁ peptide to the entire preparation. This rhythm was suppressed by MPN stimulation (firing frequency, 11 Hz), resulting in immediate termination of activity in the LG and GM neurons and the elimination of the gastric mill-timed inhibition in themvn. Shortly thereafter, DG neuron activity also terminated. The gastric mill rhythm resumed 11 sec after the end of the MPN stimulation. MPN resting potentials were −58 mV (A) and −62 mV (B). The recordings in A and B are from different preparations.

Fig. 4.

MPN increases the pyloric-timed activity of some STG gastro-pyloric neurons. A, During a time when there was an ongoing pyloric rhythm in the absence of a gastric mill rhythm, MPN stimulation (firing frequency, 13 Hz) excited the pyloric rhythm (mvn) and increased the pyloric-timed activity of Int1, VD, and IC. This is evident from the increased number of action potentials fired per pyloric-timed burst in each of these neurons. MPN stimulation also caused an increase in the amplitude of the depolarized phase of the membrane potential oscillations of each neuron, as is evident here for Int1 and in B for MG. These effects outlasted the stimulation and returned to baseline after 30 sec.B, MPN stimulation (firing frequency, 18 Hz) evoked pyloric-timed impulse activity in MG. MG began to fire action potentials that were time-locked to the pyloric rhythm during the MPN stimulation. This activity level returned to baseline after 48 sec. MPN stimulation also increased the amplitude of the membrane potential oscillations of MG. Most hyperpolarized membrane potentials: (A) Int1, −71 mV; MPN, −51 mV; (B) MG, −61 mV; MPN, −49 mV.

Fig. 5.

MPN excitation of the IC neuron elicits subthreshold pyloric-timed inhibition in the GM neuron.Left, MPN stimulation (firing frequency, 10 Hz) excited the IC and VD neurons and evoked pyloric-timed inhibition in GM. During MPN stimulation, there was also an elimination of the action potentials and tonically occurring EPSPs in GM. Right, When the IC membrane potential was hyperpolarized via DC current injection, MPN stimulation (firing frequency, 10 Hz) still excited the pyloric rhythm, but it no longer evoked pyloric-timed inhibition in GM. However, the action potentials and tonic EPSPs in GM were still eliminated. Note that when IC was hyperpolarized, its response to MPN stimulation resulted in an activity level that was slightly weaker than that occurring before MPN stimulation in the absence of hyperpolarizing current injection. Most hyperpolarized membrane potentials: (A) GM, −50 mV; IC, −58 mV; MPN, −70 mV; (B) GM, −54 mV; IC, −70 mV; MPN, −70 mV.

Fig. 6.

MPN does not inhibit the gastric mill rhythm when transmitter release is suppressed in the CoGs. A, When the CoGs were superfused with normal saline, MPN stimulation (firing frequency, 17 Hz) inhibited the gastric mill rhythm, as evident by the elimination of LG and DG bursting. B, When transmitter release was suppressed in the CoGs by superfusion with low Ca²⁺ saline, MPN stimulation (firing frequency, 17 Hz) had no effect on the gastric mill rhythm. Most hyperpolarized membrane potentials: LG, −72 mV; MPN, −50 mV. Calibration applies to both A and B.

Fig. 7.

MPN activity inhibits MCN1 and CPN2 in the CoGs.A, Left, Intracellular recordings of LG in the STG, MCN1 in the CoG and MPN in the esophageal nerve, posterior to the OG. LG is not firing action potentials but is receiving EPSPs from MCN1 and CPN2. The large-amplitude EPSPs represent input to LG from MCN1, and they are time-locked to the MCN1 action potentials. MCN1 is spontaneously active. LG is also receiving smaller amplitude EPSPs from CPN2 (see B). MPN stimulation (firing frequency, 11 Hz) inhibited MCN1, causing a cessation of MCN1 activity and hyperpolarization of its membrane potential. This eliminated all of the EPSPs in LG. Note also that the MPN stimulation excited the pyloric rhythm (mvn). Right, Schematic diagram indicating MPN inhibition of MCN1, removing excitatory input to LG.Dark cell bodies represent active neurons; light cell bodies indicate inactive neurons. Small,solid circles indicate transmitter-mediated synaptic inhibition; T-bars indicate an excitatory synapse.B, Left, Intracellular recordings of GM in the STG, CPN2 in the CoG and MPN in the esophageal nerve. GM is not firing action potentials but is receiving EPSPs from both CPN2 neurons, which are spontaneously active. MPN stimulation (firing frequency, 17 Hz) inhibited both the recorded CPN2 and its contralateral homolog, as is evident by the (1) cessation of CPN2 activity, (2) associated hyperpolarization of the CPN2 membrane potential, and (3) elimination of the EPSPs that GM was receiving. The MPN stimulation also excited the pyloric rhythm (mvn). Right, Schematic diagram indicating that MPN inhibits CPN2, removing excitatory input to GM and LG (symbols as in A). Most hyperpolarized membrane potentials: (A) LG, −44 mV; MCN1, −66 mV; MPN, −56 mV; (B) GM, −64 mV; CPN2, −56 mV; MPN, −65 mV.

Fig. 8.

MPN activity does not inhibit MCN1 or CPN2 when transmitter release in the CoGs is suppressed. A,Left, MPN stimulation (firing frequency, 14 Hz) inhibited MCN1, thereby terminating MCN1 activity and causing a hyperpolarization of the MCN1 membrane potential. Note that MPN stimulation excited the pyloric rhythm (mvn).Right, When the CoGs were selectively superfused with low Ca²⁺ saline to suppress transmitter release, MPN stimulation had no effect on MCN1. During this time, the STG was superfused with normal saline, and consequently MPN stimulation still excited the pyloric rhythm. B, Left, MPN stimulation (firing frequency, 19 Hz) inhibited CPN2, causing CPN2 to hyperpolarize and stop firing action potentials. Right, With transmitter release suppressed in the CoGs, MPN stimulation had no effect on CPN2. Most hyperpolarized membrane potentials: (A) MCN1, −65 mV; MPN, −75 mV; (B) CPN2, −63 mV; MPN, −56 mV. The recordings in A and B are from different preparations.

Fig. 9.

During an ongoing MCN1/CPN2-elicited gastric mill rhythm, MPN stimulation inhibits MCN1, CPN2, and the gastric mill rhythm. The gastric mill rhythm is represented here by the rhythmic inhibition of IC and VD (mvn) and the rhythmic bursting of the GM neuron. MCN1 and CPN2 are both firing high-frequency bursts of action potentials, and both are participating in the production of this gastric mill rhythm. CPN2 activity is monitored by an intra-axonal recording of its stomatogastric nerve axon (SNAX). MPN stimulation (firing frequency, 16 Hz) inhibited MCN1, CPN2, and the gastric mill rhythm. After MPN stimulation, MCN1 and CPN2 activity levels gradually returned to prestimulus levels, at which time the gastric mill rhythm resumed. This occurred ∼1 min after the end of the MPN stimulation. Most hyperpolarized membrane potentials: CPN2_SNAX, −66 mV; GM, −52 mV; MPN, −82 mV.

Fig. 10.

Schematic indicating the motor patterns elicited from the STG network as a result of activity in MCN1/CPN2 or MPN.Left, When MCN1 and CPN2 are active, specific versions of the pyloric rhythm (Bartos and Nusbaum, 1997) and gastric mill rhythm (Coleman and Nusbaum, 1994; Norris et al., 1994a) are elicited.Right, When MPN is active, it excites the pyloric rhythm in the STG (Nusbaum and Marder, 1989a,; present study) and inhibits MCN1 and CPN2, removing their excitation to the gastric mill system and eliminating the gastric mill rhythm (present study). This results in a distinct pyloric rhythm in the absence of the gastric mill rhythm.Dark cell bodies represent active neurons; light cell bodies represent inactive neurons.