Characterization of short-term synaptic plasticity at excitatory synapses to the supraoptic nucleus

Quinlan, Michelle (2006) Characterization of short-term synaptic plasticity at excitatory synapses to the supraoptic nucleus. Masters thesis, Memorial University of Newfoundland.

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The magnocellular neurons (MCNs) of the supraoptic nucleus (SON) synthesize and secrete the hormones oxytocin (OT) and vasopressin (AVP), which are released from the axon terminals in the neurohypophysis, as well as from the dendrites within the SON in response to physiological demand for their products. Hormone release is determined by unique firing patterns of these neurons which are dependent on intrinsic membrane properties as well as extrinsic factors such as afferent inputs. Glutamate is an important neurotransmitter in the SON, and has been shown to underlie the unique firing patterns of MCNs and to induce release of its neuropeptides. Thus, investigation of the mechanisms by which excitatory synapses to magnocellular neurons operate is critical to further our understanding of the functioning of the SON. -- In the SON, a unique form of synaptic plasticity has been shown following brief, intense stimulation to the excitatory afferents. High frequency stimulation (HFS; 10-100 Hz) causes a robust increase in the frequency and amplitude of tetrodotoxin (TTX)-insensitive miniature excitatory postsynaptic currents (mEPSCs) which lasts for 5 to 20 minutes. This has been called short term potentiation (STP). Excitability of the postsynaptic cell is increased during the course of STP, suggesting that this is a physiologically relevant phenomenon. Much is yet to be determined with reference to the time course and magnitude of STP. In particular, the larger mEPSCs that appear following HFS may increase the probability of postsynaptic firing, given that the MCNs have relatively high input resistance. However, activity-dependent potentiation of mEPSC amplitude remains poorly understood. Thus, the goal of the present thesis is to better characterize STP, with particular attention to the potentiation of mEPSC amplitude. -- Using a visually guided whole cell patch clamp recording technique, we confirmed that a 50 or 100 Hz, 1 sec stimulation of the excitatory afferents to MCN s resulted in both frequency and amplitude STP. These changes are dependent on extracellular calcium, since applying HFS in calcium free artificial cerebrospinal fluid had no effect on the amplitude or frequency of mEPSCs. First, we characterized the time course of STP. Both the amplitude and frequency of mEPSCs showed the most pronounced increase immediately following the stimulation. The initial frequency change was on average 30- 40 fold, which returned to baseline values with a two-exponential decay. The increase in amplitude was more modest compared to frequency (on average a 40% increase) which also decayed back gradually to baseline in most cells within 20 min. -- Further, we determined if variability in basal levels of spontaneous synaptic transmission seen among the cells examined explains the variability in the magnitude of the response to stimulation. We found that baseline mEPSC frequency and amplitude were negatively related to the percentage increases following HFS, which in tum were positively related to how long the mEPSC frequency and amplitude were potentiated. -- Next, we tested the hypothesis that the changes in mEPSC amplitude and frequency following HFS occur in tandem. We compared the duration of the frequency response to the duration of the amplitude response and found that they were not related: approximately half the cells tested showed longer potentiation in frequency whereas the other half had longer amplitude STP. Dissociation of these two parameters suggests that we are seeing two independent forms of synaptic plasticity. Since a similar proportion of cells showed preferential maintenance of potentiation in amplitude vs. frequency we speculated that the preference was due to the difference in phenotypes of the postsynaptic cell. However, this was not the case, as both putative oxytocin and vasopressin neurons showed both types of responses. In addition, we found that this was not a function of the intensity of stimulation, since 50 Hz and 100 Hz stimulation protocols yielded both kinds of responses. Interestingly, a high concentration of EGTA in the recording pipette to chelate postsynaptic calcium had no effect on the response of mEPSC amplitude to HFS. Addition of an NMDA receptor antagonist was also ineffective at blocking the mEPSC amplitude increase. Taken together with our observation that extracellular calcium is required for STP, these results indicate that the locus of this synaptic plasticity is presynaptic. -- From here we focused on characterizing the potentiation of mEPSC amplitude. In order to do this, we took advantage of the dissociation of the time course of mEPSC amplitude and frequency. In cells that showed potentiation of mEPSC amplitude that outlasted that of mEPSC frequency, we examined larger mEPSCs that are unlikely to have resulted from random summation of multiple mEPSCs. Analysis of mEPSC waveforms (rise and decay) revealed that the larger mEPSCs occurring when frequency has returned to baseline levels have faster rise time as compared to control. Also, in some cells, distribution histograms of mEPSC amplitude clearly showed the presence of multiple equidistant peaks following stimulation, which could indicate multiquantal transmitter release from the presynaptic terminal. -- In conclusion, this investigation illustrates the presence of two concomitant forms of synaptic plasticity arising from a HFS delivered to the excitatory synapses of the SON; one for amplitude and one for frequency of mEPSCs. Both of these forms arise from the presynaptic terminal. This result shows the capacity for sophisticated modulation of incoming excitatory information to the SON. Further investigations into the mechanisms behind these two forms of plasticity will be useful in clarifying the ways in which the excitatory afferents to the SON can modify the activity of the MCNs.

Item Type: Thesis (Masters)
Item ID: 9089
Additional Information: Includes bibliographical references (leaves 63-74).
Department(s): Medicine, Faculty of
Date: 2006
Date Type: Submission
Library of Congress Subject Heading: Neuroplasticity; Supraoptic nucleus; Synapses;
Medical Subject Heading: Neuronal Plasticity; Supraoptic Nucleus; Synapses;

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