Neurosecretion: Hypothalamic Somata versus Neurohypophysial Terminals

Abstract

The magnocellular neurons (MCNs), with their somata situated in the supraoptic and paraventricular nuclei of the hypothalamus, and their nerve terminals in the posterior pituitary (neurohypophysis), are a classical example of a neuroendocrine system. This hypothalamic-neurohypophysial system (HNS) has proven to be an important model for understanding the organization of neuronal Ca²⁺ homeostasis and mechanisms of neurosecretion. The MCNs synthesize, in a cell-specific manner, two neurohormones: arginine vasopressin (AVP) and oxytocin (OT), which can be released, in a Ca²⁺-dependent manner, both at the neurohypophysial terminal and at the somatodendritic levels. The two types of MCNs have distinct types of electrical activity leading to specific secretory patterns. OT has positive and AVP MCNs have various feedback on their own release from dendrites, but not from their axon terminals.

Action potentials and the voltage-gated Ca²⁺ channels they open are the primary regulators of [Ca²⁺]_i release in HNS terminals. Both HNS compartments utilize intracellular [Ca²⁺]_i to regulate release of their peptides. However, whereas dendrites of OT neurons utilize inositol 1,4,5-trisphosphate (IP₃) receptors, OT terminals utilize ryanodine receptors (RyRs) to regulate OT release. AVP release is not regulated in this way in either compartment. The somatodendritic AVP and OT release closely correlates with intracellular Ca²⁺ dynamics. More importantly, the Ca²⁺ stores in the endoplasmic reticulum (ER) play a major role in Ca²⁺ homeostasis in identified OT neurons. The Ca²⁺ homeostatic systems in the somata and dendrites differ from those active in the terminals; in the latter, it is mainly Ca²⁺ extrusion through the Ca²⁺ pump in the plasma membrane and uptake by mitochondria and neurosecretory granules (NSG) that are active. In both AVP and OT nerve terminals, no functional ER Ca²⁺ stores can be demonstrated experimentally. Instead, the NSG themselves store and release Ca²⁺. Nevertheless, trafficking of NSG appears to be the main mechanism for facilitation of peptide release in both compartments. Finally, SNARE-mediated exocytosis is different in HNS somata versus terminals. These fundamental differences in neurosecretion between somatodendrites and axon terminals highlight the importance of characterizing functional mechanisms in such compartments of neuroendocrine cells.

2.1 Extra Supplementary Material

Movie 2.1

Spatiotemporal dynamics of Ca²⁺ response in an isolated AVP-eGFP neuron. The spatial distribution of [Ca²⁺]_i is shown during a typical transient response induced by 50 mM K⁺. The video sequence is assembled from time lapse images of fluorescence intensity acquired video imaging of [Ca²⁺]_i was performed using an Axio Observer D1 (Zeiss) inverted microscope equipped with filters for monitoring GFP fluorescence, and epifluorescence oil immersion objectives (Plan Neofluar 100 × 1.30, FLUAR 40×/1.3 oil and FLUOR 20 × 0.75, Zeiss). This allowed us to visualize and identity the SON neurons obtained from AVP-eGFP animals. The excitation light from a Xenon lamp passed through a Lambda D4 ultrafast wavelength switching system (Sutter Instruments) with a maximum switching frequency of 500 Hz. The fluorescence intensity was detected by using a cooled CCD camera (AxioCam MRm, Zeiss) and the whole system was controlled by Zeiss ZEN Imaging software (2012-SP2/AxioVision SE64 Rel. 4.8.3). The fluorescence intensity was measured with excitations at 340 and 380 nm, and emission at 510 nm. The color scale corresponds to the estimated local [Ca²⁺]_i value. The elapsed time is shown at bottom left. The integrated [Ca²⁺]_i value is given at bottom left and in the progressively drawn trace, with green bar indicating the time interval during which K⁺ was applied. Modified from Kortus et al. (2016a) with Courtesy of “Science Direct” (MP4 3411 kb)

Movie 2.2

Spatiotemporal dynamics of spontaneous Ca²⁺ oscillations in an isolated AVP-eGFP neuron. As described above, the spatial distribution of [Ca²⁺]_i is shown during a spontaneous oscillation in normal condition. The video sequence is assembled from time lapse images of fluorescence intensity, acquired as described above. The color scale corresponds to the estimated local [Ca²⁺]_i value. The elapsed time is shown at top left. The integrated [Ca²⁺]_i value is given at top left and in the progressively drawn trace. Modified from Kortus et al. (2016a) with Courtesy of “Science Direct” (MP4 11701 kb)

Movie 2.3

Intracellular calcium sparks in NH terminals. Movie of murine HNS terminal patched in whole-cell configuration and loaded with Fluo-3 to visualize intracellular calcium. Calcium free bath solution, thus any calcium must come from internal stores. Note the number of calcium sparks or “syntillas” which appear spontaneously. Depolarizations, however, increase the number but not the size of such intraterminal calcium release events. (Courtesy of Dr. Valerie DeCrescenzo and Biomedical Imaging Group) (WMV 281 kb)

Movie 2.4

Movie Mechanisms for release facilitation in neurohypophysial terminals (NHT). Neurosecretory granule (NSG) and microvesicle (MV) exocytosis is known to be a Ca²⁺-dependent process. Release of a significant amount of Ca²⁺ precisely where exocytosis occurs could conceivably affect release (1). Alternatively, ryanodine-sensitive (RyR) Ca²⁺ stores could functionally modulate the size of vesicular release pools in NHT (2). The voltage dependence of syntillas, through their coupling to L-type Ca²⁺ channels (DHPR), would appear to make them ideal candidates to drive such a mechanism. Such a mechanism could serve as an activity-dependent means to recruit NSG from reserve pools in the cytosol, to the readily releasable pool (RRP) and immediately releasable pool (IRP) adjacent to the plasma membrane. (Courtesy of Dr. James McNally) (AVI 30345 kb)

Key References: See Main List for Reference Details

• Cazalis et al. (1987) First demonstration of depolarization–secretion coupling (DSC) in isolated terminals from the hypothalamic-neurohypophysial system (HNS).

• Chow et al. (1996) First demonstration of fusion pore opening before full secretion.

• Dayanithi et al. (1996) First demonstration to show that in both AVP and OT neurons, all calcium channels are involved in high K⁺-induced [Ca²⁺]_i responses but AVP-induced [Ca²⁺]_i responses are mostly activated by L, N, P/Q, and R-type channels, whereas, OT-induced [Ca²⁺]_i responses are exclusively from the release of Ca²⁺ from intracellular IP₃/TG-sensitive Ca²⁺ stores.

• Douglas and Poisner (1964) Review of calcium as the coupler in depolarization–secretion coupling (DSC).

• Giovannucci and Stuenkel (1997) Demonstration that calcium regulates trafficking of granules between different secretory pools.

• Gouzenes et al. (1999a, b) First demonstration that not only AVP-V_1a receptors but also V₂-type (classically restricted in peripheral system-PNS) AVP receptors regulate AVP-induced [Ca²⁺]_i responses in the CNS (central nervous system) neurons. We assume that the AVP receptory sub-types expressed in the PNS and CNS might have different pharmacological profiles depending on the physiological status of the animal.

• Jin et al. (2007) First demonstration that ryanodine receptors play critical role in oxytocin release.

• Kortus et al. (2016a, b) This is the first detailed demonstration to clearly show the spontaneous [Ca²⁺]_i oscillations in the isolated AVP and OT neurons from the transgenic rats for AVP-eGFP and OT-mRFP.

• Ludwig et al. (2002) Demonstration that neuropeptide release from somatodendrites can be facilitated by intracellular calcium.

• McNally et al. (2014) First demonstration that neurosecretory granules (NSG) can serve as functional calcium stores in nerve terminals.

• Zhang and Zhou (2002) First demonstration of Ca²⁺-independent but voltage-dependent secretion (CIVDS) in neuronal cell bodies.