Incremental Temperature Changes for Maximal Breeding and Spawning in Astyanax mexicanus

Summary

This article outlines the basic laboratory conditions and protocols for an incremental temperature regime to stimulate maximal spawning in the Mexican tetra Astyanax mexicanus, which is an emerging model for developmental and evolutionary studies.

Abstract

The Mexican tetra, Astyanax mexicanus, is an emerging model system for studies in development and evolution. The existence of eyed surface (surface fish) and blind cave (cave fish) morphs in this species presents an opportunity to interrogate the mechanisms underlying morphological and behavioral evolution. Cave fish have evolved novel constructive and regressive traits. The constructive changes include increases in taste buds and jaws, lateral line sensory organs, and body fat. The regressive changes include loss or reduction of eyes. melanin pigmentation, schooling behavior, aggression, and sleep. To experimentally interrogate these changes, it is crucial to obtain large numbers of spawned embryos. Since the original A. mexicanus surface fish and cave fish were collected in Texas and Mexico in the 1990s, their descendants have been routinely stimulated to breed and spawn large numbers of embryos bimonthly in the Jeffery laboratory. Although breeding is controlled by food abundance and quality, light-dark cycles, and temperature, we have found that incremental temperature changes play a key role in stimulating maximal spawning. The gradual increase of temperature from 72 °F to 78 °F in the first three days of a breeding week provides two-three consecutive spawning days with maximal numbers of high-quality embryos, which is then followed by a gradual decrease of temperature from 78 °F to 72 °F during the last three days of the spawning week. The procedures shown in this video outline the workflow before and during a laboratory breeding week for incremental temperature stimulated spawning.

Introduction

The teleost Astyanax mexicanus has an eyed surface-dwelling (surface fish) form and many different blind cave-dwelling (cave fish) forms^1,2. Cave fish have evolved in perpetual darkness and under food limitations, resulting in the appearance of novel constructive and regressive traits³. The constructive traits include increases in taste buds and jaw size, sensory organs of the lateral line, and fat reserves. The regressive traits include the loss or reduction of melanin pigmentation, eyes, and behaviors, such as sleep, schooling, and aggression. An attribute of the Astyanax....

Protocol

This procedure has been approved by Institutional Animal Care guidelines of the University of Maryland, College Park (Currently IACUC 469 #R-NOV-18-59; Project 1241065-1).

Figure 1. Calendars during a breeding week and a non-breeding week. Please click here....

Results

We generally breed and spawn the descendants of surface fish originally collected at Nacimiento del Rio Choy in San Luis Potosi, Mexico (Rio Choy surface fish) and San Solomon Springs in Balmorhea State Park, Texas (Texas surface fish) and cave fish derived from Cueva de El Pachón (Pachón cave fish) in Tamaulipas, Mexico, and Cueva de los Sabinos (Los Sabinos cave fish) and Sotano de la Tinaja (Tinaja cave fish) in San Luis Potosi, Mexico.  

Throughout a breeding week, data is collected for va.......

Discussion

Astyanax mexicanus is a novel biological model that spawns frequently and can be bred easily in the laboratory^1,2. Because we are interested in the developmental mechanisms that underlie evolutionary changes in A. mexicanus cave fish, the production and use of embryos is vital to our research goals. The primary purpose of maintaining an adult stock of fish is the production of embryos and young fry for use in developmental experiments and for re.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Blackworms	Eastern Aquatics, Lancaster, PA	None
Breeding Nets	Custom made
Brine shrimp eggs	AquaCave Lake Forest, IL.	None
Colorimetric test kit	Petco	SKU:11916	API Freshwater pH Test Kit
Egg yolk flakes	Pentair, Minneapolis, MN	None
Fingerbowls	Carolina Biological Supply	741004	Culture dishes, 4.5 in, 250 mL
Hand held nets	Any Pet Store
Incubator for embryos	Fisher Scientific	51-029-321HPM	405 L
Instant Ocean sea salts	Spectrum Brands, Blacksburg, VA	None
Methylene Blue	Sigma-Aldrich, St. Louis, MO	M9140
Pasteur Pipettes	Fisher Scientific	13-678-20	5.75 in.
Net soaking solution	Any Pet Store
Nutrafin Cycle	Amazon	None	Bacterial boost
Refrigerator for live feed	Any source
Stereomicroscope	Any source
Thermometer	Any source
Tetra Tropical Crisps	Spectrum Brands, Blacksburg, VA	None