command neurons in tt'? edible snail (Helix pomatia, Helix lucorum) after training to a conditioned defensive reflex (CDR). Prior to experiments, snails were kept ...
Neuroscience and Behavioral Physiology, Vol. 27, No. 4, 1997
CHANGES IN THE ELECTRICAL CHARACTERISTICS OF COMMAND NEURONS ON DEVELOPMENT
OF A
CONDI-TIONED DEFENSIVE REFLEX IN THE EDIBLE SNAIL
Kh. L. Gainutdinov, T. Kh. Gainutdinova, and L. Yu. Chekmarev
UDC 612.822.3 + 612.821.6
The role of excitability in the phenomenon of plasticity is presently subject to intense study [2-4]. However, most investigations are carried out using simplified models in which the mechanisms of plasticity, which are the basis of associative learning, are not analyzed. The aim of the present work was to study the electrical characteristics of defensive behavior command neurons in tt'? edible snail (Helix pomatia, Helix lucorum) after training to a conditioned defensive reflex (CDR). Prior to experiments, snails were kept for at least two weeks in an active state in a glass terrarium with a humid atmosphere at room temperature. A CDR was developed as described in [2]. The conditioned stimulus was a tap on the shell, which in normal conditions does not produce a defensive response in the snails. The unconditioned stimulus consisted of directing a burst of air into the pulmonary cavity, which induced an unconditioned defensive response consisting of closure of the pneumostoma. Combinations of stimuli were presented at intervals of 2-4 min; the reflex was developed by an accelerated method over 3-4 days, after presentation of 140-160 combinations. This training procedure induced complete closure of the pneumostoma in response to the conditioned stimulus, which was taken as a positive response. Snails in the active control group were presented with the same number of conditioned and unconditioned stimuli as used in the experimental group, but in random order. Positive responses in the active controi group amounted to 25 %, which was significantly lower than the level in the experimental snails. After training, the electrical characteristics of the following defensive behavior command neurons were recorded: LPa3, PPa3, LPa2, and PPa2. Measurements were made using intracellular glass microelectrodes with a resistance of 5-25 MfL During experiments, recordings were made of the following parameters of neuron electrical activity: resting potential, action potential amplitude, and action potential generation threshold. Measurements were carried out using isolated snail central nervous systems (n = 51): 25 from control animals, 18 after development of the CDR, and 8 active control snails. Totals of 43 LPa3 and PPa3 neurons and 22 LPa2 and PPa2 neurons were studied. Results were analyzed statistically using Student's t test and the W coefficient of Wilcoxon for linked pairs. Studies of the electrical characteristics of command neurons showed that the membrane potential of LPa3 and PPa3 neurons reached values of - 6 2 . 7 + 1 mV, and the action potential generation threshold was 20.8 + 0.4 mV. These parameters had similar values for LPa2 and PPa2 neurons (Tables 1 and 2). In snails which had developed the CDR, command neurons LPa3, PPa3, LPa2, and PPa2 showed significant (p < 0.05) reductions in the membrane potential (Table 1), both for neurons LPa3 and PPa3 (n = 22) and for neurons LPa2 and PPa2 (n = 8). Additionally, there were significant (p < 0.05) reductions in the action potential generation thresholds (Table 2). Results obtained in active controls (n < 9) showed only small differences from the electrical characteristics of command neurons in intact snails (Tables 1 and 2). Measurements of action potential amplitudes showed that this parameter underwent no significant change on development of the reflex. The electrical characteristics of motor neurons involved in opening and closing of the pneumostoma in intact snails, in the active control group, and in the experimental series, were not significantly different. These results show that associative learning is accompanied by significant changes in the membrane potential in defensive behavior command neurons. This is primarily a depolarization shift of the initial resting potential of the cells by 5-6 Kazan' Physicotechnical Institute, Cardiological Scientific Center, Russian Academy of Sciences, Kazan' State University. Translated from Zhurnal Vysshei Nervnoi Deyatel'nosti, Vol. 46, No. 3, pp. 614-616, May-June, 1996. Original article submitted January 9, 1996; accepted for publication February 22, 1996. 0097-0549/97/2704-0367518.00 9
Plenum Publishing Corporation
367
TABLE 1. Membrane Potentials (mV) in Command Neurons LPa3, PPa3, LPa2, and PPa2 during Development of a Conditioned Defensive Reflex (CDR) in Intact Animals (C) and Active Controls (AC) Group of neurons LPa3 and PPa3 LPa2 and PPa2
C
CDR
AC
- - 62,7 4- 1,0 - - 61,5 4- !,5
-- 57,2 4- 1,0 -- 56,9.4- 1,1
- - 61,4 -+- 1,7 - - 61,4 -4- 1,7
TABLE 2. Threshold Potentials (mV) of Command Neurons LPa3, PPa3, LPa2, and PPa2 during Development of a Conditioned Defensive Reflex (CDR) in Intact Animals (C) and Active Controls (AC) Group of neurons LPa3 and PPa3 LPa2 and PPa2
C
CDR
AC
20,8 -4- 0,4 19,6 4- 0,4
16,1 4- 0,4 16,5 -4- 0,4
19,8 -4- 0,6 19,8 -4- 0,6
mV, with a reduction in the action potential generation threshold by some 4 mV; this indicates an increase in the excitability of command neurons on development of the conditioned reflex. A similar result was obtained previously on development of the same conditioned reflex in semi-intact preparations. In this case, increases were seen in the excitability of command neurons LPa3 and PPa3, which control the defensive behavior consisting of pneumostoma closure in edible snails. At the early stages of CDR development, spontaneous action potentials appeared by 25-30 combinations of the conditioned and unconditioned stimuli [1]; additionally, large numbers of action potentials were generated after development of the conditioned reflex [2]. In Aplysia, Kandel's group found [5] that associative learning was accompanied by an increase in the amplitude of the excitatory post-synaptic potential from the sensory neuron to motor neuron L7, due to reduced efflux potassium currents and increased influx calcium currents. During development of the conditioned reflex, the slow inactivation of the calcium current creates conditions in which stimuli arriving at different times can be associated. In addition, during training of Hermissenda, longlasting reductions in amplitude were seen, along with increases in the rate of inactivation, in rapidly inactivating potassium currents of type B photoreceptors [4]. Comparison of the arcs of the unconditioned and conditioned defensive reflexes suggests that these are closed at the level of command and sensory defensive reflex neurons, as well as in the sensory and interneuron apparatus involved in spatial perception, which receives the conditioned stimulus [2, 3, 5]. This leads to the idea that this modification of the state of the nervous system has the effect that sensory and command neurons become memory cells and, simultaneously, functional elements fulfilling one or another function, depending on the parameters of the external signals. These results are in agreement with many data indicating that command and sensory neurons are the plastic elements. These data, along with results obtained by other authors [2], suggest that motor neurons are not plastic elements in these types of modification of the state of the nervous system. The universal concept, of changes in the efficiency of excitatory synapses, is particularly indicated for nonassociative forms of learning and associative strengthening of defensive responses in invertebrates. Thus, these results demonstrate an increase in excitability in defensive behavior command neurons during associative learning in animals, expressed as a reduction in the action potential generation threshold. The authors would like to thank A. E. Tarasov for assistance with these studies. These investigations were supported by the Russian Fund for Basic Research (grant No. 95-04-12603).
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E . G . Litvinov and D. B. Logunov, "Changes in the excitability of command neurons in the initial period of formation of a conditioned reflex in the edible snail," Zh. Vyssh. Nerv. Deyat., 29, No. 2, 284 (1979).
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